McGuigan, K G; Méndez-Hermida, F; Castro-Hermida, J A; Ares-Mazás, E; Kehoe, S C; Boyle, M; Sichel, C; Fernández-Ibáñez, P; Meyer, B P; Ramalingham, S; Meyer, E A
To determine whether batch solar disinfection (SODIS) can be used to inactivate oocysts of Cryptosporidium parvum and cysts of Giardia muris in experimentally contaminated water. Suspensions of oocysts and cysts were exposed to simulated global solar irradiation of 830 W m(-2) for different exposure times at a constant temperature of 40 degrees C. Infectivity tests were carried out using CD-1 suckling mice in the Cryptosporidium experiments and newly weaned CD-1 mice in the Giardia experiments. Exposure times of > or =10 h (total optical dose c. 30 kJ) rendered C. parvum oocysts noninfective. Giardia muris cysts were rendered completely noninfective within 4 h (total optical dose >12 kJ). Scanning electron microscopy and viability (4',6-diamidino-2-phenylindole/propidium iodide fluorogenic dyes and excystation) studies on oocysts of C. parvum suggest that inactivation is caused by damage to the oocyst wall. Results show that cysts of G. muris and oocysts of C. parvum are rendered completely noninfective after batch SODIS exposures of 4 and 10 h (respectively) and is also likely to be effective against waterborne cysts of Giardia lamblia. These results demonstrate that SODIS is an appropriate household water treatment technology for use as an emergency intervention in aftermath of natural or man-made disasters against not only bacterial but also protozoan pathogens.
Craik, Stephen A; Smith, Daniel W; Chandrakanth, Mysore; Belosevic, Miodrag
Static mixers may be used to dissolve gaseous ozone in water treatment facilities in order to provide protection against the waterborne parasite Cryptosporidium parvum. The objective of this study was to determine the effect of a brief exposure to turbulent gas-liquid mixing conditions in a static mixer on inactivation of C. parvum oocysts by ozone. Inactivation measured in an ozone contacting apparatus that employed a static mixer for ozone dissolution was compared to predictions based on a previously published kinetic model of C. parvum inactivation by dissolved ozone in gently stirred batch reactors. Although initial contact in the static mixer had no immediate effect on the oocysts, a 20% increase in the rate of inactivation during subsequent contact with dissolved ozone was observed. Increasing the degree of turbulence within the static mixer did not yield additional inactivation. Use of static mixers for dissolution of ozone in drinking water treatment systems may provide limited enhancement of C. parvum inactivation by dissolved ozone.
This research was conducted to determine the prevalence of Cryptosporidium oocysts among children with acute gastroenteritis in Zaria, Nigeria by Kinyoun Modified Carbol-Fuchsin Staining (Modified Ziehl-Neelsen Staining) Technique. The results for the screening of Cryptosporidium oocysts showed that out of 372 stool ...
Nichols, Rosely A B; Campbell, Brian M; Smith, Huw V
We developed and validated a PCR-based method for identifying Cryptosporidium species and/or genotypes present on oocyst-positive microscope slides. The method involves removing coverslips and oocysts from previously examined slides followed by DNA extraction. We tested four loci, the 18S rRNA gene (N18SDIAG and N18SXIAO), the Cryptosporidium oocyst wall protein (COWP) gene (STN-COWP), and the dihydrofolate reductase (dhfr) gene (by multiplex allele-specific PCR), for amplifying DNA from low densities of Cryptosporidium parvum oocysts experimentally seeded onto microscope slides. The N18SDIAG locus performed consistently better than the other three tested. Purified oocysts from humans infected with C. felis, C. hominis, and C. parvum and commercially purchased C. muris were used to determine the sensitivities of three loci (N18SDIAG, STN-COWP, and N18SXIAO) to detect low oocyst densities. The N18SDIAG primers provided the greatest number of positive results, followed by the N18SXIAO primers and then the STN-COWP primers. Some oocyst-positive slides failed to generate a PCR product at any of the loci tested, but the limit of sensitivity is not entirely based on oocyst number. Sixteen of 33 environmental water monitoring Cryptosporidium slides tested (oocyst numbers ranging from 1 to 130) contained mixed Cryptosporidium species. The species/genotypes most commonly found were C. muris or C. andersoni, C. hominis or C. parvum, and C. meleagridis or Cryptosporidium sp. cervine, ferret, and mouse genotypes. Oocysts on one slide contained Cryptosporidium muskrat genotype II DNA.
Petersen, Heidi Huus; Enemark, Heidi L.
Viability assessment of Cryptosporidium parvum oocysts is crucial for evaluation of the public health significance of this important zoonotic protozoon. Viability is commonly assessed in wet mounts after acid pretreatmentand staining with fluorogenic vital dyes. However, in some studies, oocyst...
Liu, Yuanyuan; Zhang, Changyong; Hilpert, Markus; Kuhlenschmidt, Mark S.; Kuhlenschmidt, Theresa B.; Nguyen, Thanh H.
Effective removal of Cryptosporidium parvum oocysts by granular filtration requires the knowledge of oocyst transport and deposition mechanisms, which can be obtained based on real time microscopic observation of oocyst transport in porous media. Attachment of oocysts to silica surface in a radial stagnation point flow (RSPF) cell and in a micromodel, which has 2-dimensional (2-D) microscopic pore structures consisting of an array of cylindrical collectors, was studied and compared. Real time transport of oocysts in the micromodel was recorded to determine the attached oocyst distributions in transversal and longitudinal directions. In the micromodel, oocysts attached to the forward portion of clean collectors, where the flow velocity was lowest. After initial attachment, oocysts attached onto already attached oocysts. As a result, the collectors ripened and the region available for flow was reduced. Results of attachment and detachment experiments suggest that surface charge heterogeneity allowed for oocyst attachment. In addition to experiments, Lattice-Boltzmann simulations helped understanding the slightly non-uniform flow field and explained differences in the removal efficiency in the transversal direction. However, the hydrodynamic modeling could not explain differences in attachment in the longitudinal direction.
Hønsvall, Birgitte K; Robertson, Lucy J
Both Cryptosporidium parvum and Cryptosporidium hominis are often associated with cryptosporidiosis in humans, but whereas humans are the main host for C. hominis, C. parvum is zoonotic and able to infect a variety of species. The oocyst transmission stages of both species of parasites are morphologically identical and molecular techniques, usually polymerase chain reaction (PCR), are required to distinguish between oocysts detected by standard methods in environmental samples, such as water. In this study, we developed two primer sets for real-time nucleic acid sequence-based amplification (NASBA), targeting the MIC1 transcript in C. parvum (CpMIC1) and C. hominis (ChMIC1). Using these primer sets, we were not only able to detect low numbers of C. parvum and C. hominis oocysts (down to 5 oocysts in 10 μl, and down to 1 oocyst using diluted RNA samples), but also distinguish between them. One of the primer sets targeted an exon only occurring in CpMIC1, thereby providing a tool for distinguishing C. parvum from other Cryptosporidium species. Although mRNA has been suggested as a tool for assessing viability of Cryptosporidium oocysts, as it is short-lived and may have high transcription, this NASBA assay detected MIC1 mRNA in inactivated oocysts. RNA within the oocysts seems to be protected from degradation, even when the oocysts have been killed by heating or freeze-thawing. Thus, our approach detects both viable and non-viable oocysts, and RNA does not seem to be a suitable marker for assessing oocyst viability. Copyright © 2016 Elsevier Inc. All rights reserved.
associated with feacal pollution does not necessarily indicate the absence of Cryptosporidium or Giardia. Being a common cause of waterborne disease, the requirement for detecting the presence of these organisms in environmental and treated water samples is growing. Although cysts and oocysts can be found in natural ...
The present study was conducted to investigate the point prevalence of Cryptosporidium oocysts infection in calves grazing along the bank of Rima River Sokoto in October 2011. The river bank is a converging zone for domestic animals reared in different quarters of the town and the surrounding settlements. A total number ...
Bukhari, Z; Smith, H V
Bovine fecal samples (1 g) negative for Cryptosporidium sp. oocysts were seeded with 7 x 10(4) Cryptosporidium parvum oocysts and purified by either water-ether concentration, sucrose density flotation, or zinc sulfate flotation to evaluate oocyst recovery. The effect of these purification techniques on the viability of recovered oocysts was also evaluated. Significantly higher numbers of seeded oocysts were recovered by water-ether concentration (recovery rate, 46 to 75%) than by sucrose den...
Li, S.Y.; Goodrich, J.A.; Owens, J.H. [Environmental Protection Agency, Cincinnati, OH (United States)] [and others
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.
Cryptosporidium is a chlorination-resistant protozoan parasite that causes a self-limiting diarrheal disease in the immunocompetent or severe chronic diarrhea in the immunocompromised. Two species, C. parvum and C. hominis, cause most cases of cryptosporidiosis in humans, while C...
Xiao, Lihua; Singh, Ajaib; Limor, Josef; Graczyk, Thaddeus K.; Gradus, Steve; Lal, Altaf
Recent molecular characterizations of Cryptosporidium parasites make it possible to differentiate the human-pathogenic Cryptosporidium parasites from those that do not infect humans and to track the source of Cryptosporidium oocyst contamination in the environment. In this study, we used a small-subunit rRNA-based PCR-restriction fragment length polymorphism (RFLP) technique to detect and characterize Cryptosporidium oocysts in 55 samples of raw surface water collected from several areas in t...
Cheng, Hui-Wen A; Lucy, Frances E; Graczyk, Thaddeus K; Broaders, Michael A; Tamang, Leena; Connolly, Michelle
This study investigates the fate of Cryptosporidium parvum and C. hominis oocysts and Giardia duodenalis cysts at four Irish municipal wastewater treatment plants (i.e., Plant A, B, C, and D) that utilize sludge activation or biofilm-coated percolating filter systems for secondary wastewater treatment. The fate of these pathogens through the sewage treatment processes was determined based on their viable transmissive stages, i.e., oocysts for Cryptosporidium and cysts for Giardia. Analysis of final effluent indicated that over 97% of viable oocysts and cysts were eliminated, except at Plant C, which achieved only 64% of oocyst removal. A significant correlation between the removal of oocysts and cysts was found at Plants A, B, and D (R = 0.98, P hominis, and G. duodenalis, with maximum concentrations of 20 oocysts and eight cysts per gram in primary sludge indicating the need for further sludge sanitization treatments. This study provides evidence that C. parvum and C. hominis oocysts and G. duodenalis cysts are present throughout the wastewater processes and in end-products, and can enter the aquatic environment with consequent negative implications for public health.
Al-Adhami, B.H.; Nichols, R.A.B.; Kusel, J.R.; O'Grady, J.; Smith, H.V.
To investigate the effect of UV light on Cryptosporidium parvum and Cryptosporidium hominis oocysts in vitro, we exposed intact oocysts to 4-, 10-, 20-, and 40-mJ·cm−2 doses of UV irradiation. Thymine dimers were detected by immunofluorescence microscopy using a monoclonal antibody against cyclobutyl thymine dimers (anti-TDmAb). Dimer-specific fluorescence within sporozoite nuclei was confirmed by colocalization with the nuclear fluorogen 4′,6′-diamidino-2-phenylindole (DAPI). Oocyst walls we...
Gómez-Couso, Hipólito; Fontán-Sainz, María; Ares-Mazás, Elvira
To determine the thermal contribution, independent of ultraviolet radiation, on the inactivation of Cryptosporidium parvum during solar water disinfection procedures (SODIS), oocysts were exposed for 4, 8, and 12 hours to temperatures recorded in polyethylene terephthalate bottles in previous SODIS studies carried out under field conditions. Inclusion/exclusion of the fluorogenic vital dye propidium iodide, spontaneous excystation, and infectivity studies were used to determine the inactivation of oocysts. There was a significant increase in the percentage of oocysts that took up propidium iodide and in the number of oocysts that excysted spontaneously. There was also a significant decrease in the intensity of infection elicited in suckling mice at the end of all exposure times. The results of the study demonstrate the importance of temperature in the inactivation of C. parvum oocysts during application of SODIS under natural conditions.
Ono, Kazuo; Tsuji, Hidetaka; Rai, Shiba Kumar; Yamamoto, Akio; Masuda, Kuniyoshi; Endo, Takuro; Hotta, Hak; Kawamura, Takashi; Uga, Shoji
In Japan, only a few rivers have been inspected for Cryptosporidium parvum contamination, and the methods used had low sensitivity. In 1998 and 1999, we used a method with higher sensitivity to examine all large rivers used as sources of water supply in one prefecture (which we divided into four areas) in western Japan for Cryptosporidium oocysts. One sample was collected at each of 156 sites along 18 rivers, and samples were tested for Cryptosporidium oocysts by immunomagnetic separation. Sa...
Al-Sabi, Mohammad Nafi Solaiman; Gad, J. A.; Riber, Ulla
AimsTo develop a filtration unit for efficient recovery of waterborne Cryptosporidium oocysts and Giardia cysts ((oo-)cysts) in drinking water.Methods and ResultsThis unit utilizes a metallic filter and an ultrasound transducer for eluting (oo-)cysts, with a fixed retentate backwash volume; approx...... was assessed by seeding ColorSeed (10 replicates) into the filter unit followed by air backwash to a glass slide and counting of (oo-)cysts by epifluorescent microscopy. High recovery rates (meanSD) were found: 849%48 for Giardia cysts and 70%+/- 65 for Cryptosporidium oocysts. DNA of seeded wild type (oo...
Bukhari, Z; Smith, H V
Bovine fecal samples (1 g) negative for Cryptosporidium sp. oocysts were seeded with 7 x 10(4) Cryptosporidium parvum oocysts and purified by either water-ether concentration, sucrose density flotation, or zinc sulfate flotation to evaluate oocyst recovery. The effect of these purification techniques on the viability of recovered oocysts was also evaluated. Significantly higher numbers of seeded oocysts were recovered by water-ether concentration (recovery rate, 46 to 75%) than by sucrose density (24 to 65%) or zinc sulfate (22 to 41%) flotation methods. In addition, water-ether concentration did not exert a significant effect on the viability of the population of oocysts recovered, whereas sucrose density flotation and zinc sulfate flotation selectively concentrated viable oocysts. The water-ether concentration procedure is recommended for use in epidemiological studies in which both oocyst enumeration and viability assessment are required. PMID:8567888
Solo-Gabriele Helena María
Full Text Available During June 1996, water supplies of the city of San Pedro Sula, Honduras, were sampled to obtain an assessment of Cryptosporidium oocyst and Giardia cyst concentrations. Each sample was concentrated and stained with an indirect immunofluorescent antibody, and parasites were counted through microscopic analysis. In three surface water supplies, Cryptosporidium oocyst concentrations ranged from 58 to 260 oocysts per 100 L, and Giardia cysts were present in concentrations ranging from 380 to 2100 cysts per 100 L. Unlike the surface water samples, groundwater had a higher concentration of Cryptosporidium oocysts (26/100 L than Giardia cysts (6/100 L, suggesting that the groundwater aquifer protects the water supply more effectively from larger Giardia cysts. Cryptosporidium oocyst concentrations are within the typical range for surface water supplies in North America whereas Giardia cyst concentrations are elevated. Efforts should be made to protect raw water from sources of contamination.
Power, Michelle L; Sangster, Nicholas C; Slade, Martin B; Veal, Duncan A
The occurrence of Cryptosporidium oocysts in feces from a population of wild eastern grey kangaroos inhabiting a protected watershed in Sydney, Australia, was investigated. Over a 2-year period, Cryptosporidium oocysts were detected in 239 of the 3,557 (6.7%) eastern grey kangaroo fecal samples tested by using a combined immunomagnetic separation and flow cytometric technique. The prevalence of Cryptosporidium in this host population was estimated to range from 0.32% to 28.5%, with peaks occurring during the autumn months. Oocyst shedding intensity ranged from below 20 oocysts/g feces to 2.0 x 10(6) oocysts/g feces, and shedding did not appear to be associated with diarrhea. Although morphologically similar to the human-infective Cryptosporidium hominis and the Cryptosporidium parvum "bovine" genotype oocysts, the oocysts isolated from kangaroo feces were identified as the Cryptosporidium "marsupial" genotype I or "marsupial" genotype II. Kangaroos are the predominant large mammal inhabiting Australian watersheds and are potentially a significant source of Cryptosporidium contamination of drinking water reservoirs. However, this host population was predominantly shedding the marsupial-derived genotypes, which to date have been identified only in marsupial host species.
Johnson, Anne M.; Linden, Karl; Ciociola, Kristina M.; De Leon, Ricardo; Widmer, Giovanni; Rochelle, Paul A.
The Cryptosporidium spp. UV disinfection studies conducted to date have used Cryptosporidium parvum oocysts. However, Cryptosporidium hominis predominates in human cryptosporidiosis infections, so there is a critical need to assess the efficacy of UV disinfection of C. hominis. This study utilized cell culture-based methods to demonstrate that C. hominis oocysts displayed similar levels of infectivity and had the same sensitivity to UV light as C. parvum. Therefore, the water industry can be ...
EPA Methods 1622 and 1623 are the benchmarks for detection of Cryptosporidium spp. oocysts in water. These methods consist of filtration, elution, purification by immunomagnetic separation (IMS), and microscopic analysis after staining with a fluorescein isothiocyanate conjugate...
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...
Schets FM; Medema GJ; MGB
The incorporation of flow cytometry as an additional purification step has improved the detection method for Cryptosporidium oocysts and Giardia cysts in water. Flow cytometry allows separation of (oo)cysts from interfering debris particles present in water concentrates and thus enables the
The ability of infectious oocyst forms of Toxoplasma gondii and Cryptosporidium spp. to resist disinfection treatments and cause disease may have significant public health implications. Currently, little is known about oocyst-specific factors involved during host cell invasion p...
The ability of infectious oocyst forms of Toxoplasma gondii and Cryptosporidium spp. to resist disinfection treatments and cause disease may have significant public health implications. Currently, little is known about oocyst-specific factors involved during host cell invasion pr...
Gómez-Couso, H; Fontán-Saínz, M; Sichel, C; Fernández-Ibáñez, P; Ares-Mazás, E
To investigate the efficacy of the solar water disinfection (SODIS) method for inactivating Cryptosporidium parvum oocysts in turbid waters using 1.5 l polyethylene terephthalate (PET) bottles under natural sunlight. All experiments were performed at the Plataforma Solar de Almería, located in the Tabernas Desert (Southern Spain) in July and October 2007. Turbid water samples [5, 100 and 300 nephelometric turbidity units (NTU)] were prepared by addition of red soil to distilled water, and then spiked with purified C. parvum oocysts. PET bottles containing the contaminated turbid waters were exposed to full sunlight for 4, 8 and 12 h. The samples were then concentrated by filtration and the oocyst viability was determined by inclusion/exclusion of the fluorogenic vital dye propidium iodide. Results After an exposure time of 12 h (cumulative global dose of 28.28 MJ/m(2); cumulative UV dose of 1037.06 kJ/m(2)) the oocyst viabilities were 11.54%, 25.96%, 41.50% and 52.80% for turbidity levels of 0, 5, 100 and 300 NTU, respectively, being significantly lower than the viability of the initial isolate (P < 0.01). SODIS method significantly reduced the potential viability of C. parvum oocysts on increasing the percentage of oocysts that took up the dye PI (indicator of cell wall integrity), although longer exposure periods appear to be required than those established for the bacterial pathogens usually tested in SODIS assays. SODIS.
Graczyk, T. K.; Fayer, R.; Cranfield, M. R.; Conn, D. B.
Corbicula fluminea hemocytes phagocytosed infectious oocysts of Cryptosporidium parvum in vitro. After 15, 30, 60, 90, and 120 min of incubation, averages of 35.8, 58.0, 69.7, 77.7, and 81.6% of the oocysts were phagocytosed by 24.3, 70.0, 78.5, 87.3, and 93.0% of the hemocytes, respectively. A single clam can retain by phagocytosis an average of 1.84 x 10(sup6) oocysts per ml of hemolymph. C. fluminea bivalves can serve as biological indicators of contamination of wastewaters and agricultural drainages with Cryptosporidium. PMID:16535656
R. G. Mohammad
The study included examination of 110 water samples from well distributed in Mosul city and few towns and villages around it from May 2009 to March 2010 for detection of Cryptosporidium spp oocysts and Giardia spp cysts in well water. The results revealed that the prevalence of Cryptosporidium oocysts was 16.36% while the prevalence of Giardia cysts was 12.72%. The percentages of prevalence with Cryptosporidium and Giardia were in high rate in Bartilla and some villages around it 20% for Cryp...
Osaki, Silvia Cristina; Costa, Adriana Oliveira; Troiano, Ludmilla Della Coletta; Kruger, Ernesto Renato; Pereira, Juliana Tracz; Fernandes, Nelson Luis Mello; Silva, Márcia Benedita de Oliveira; Soccol, Vanete Thomaz
The production of anti-Cryptosporidium polyclonal antibodies and its use in direct immunofluorescence assays to determine the presence of Cryptosporidium in water are described in the present work. Two rabbits were immunized with soluble and particulate antigens from purified Cryptosporidium oocysts. The sera produced were prepared for immunoglobulin G extraction, which were then purified and conjugated with fluorescein isothiocyanate (FITC). Slides containing known amounts of oocysts were prepared to determine the sensitivity of the technique. To test the specificity, slides containing Giardia duodenalis cysts were prepared. The conjugate was successfully used in water samples experimentally contaminated with Cryptosporidium oocysts, and it was possible to detect up to five oocysts/spot, corresponding to contamination of 250 oocysts/mL. The three immunizations performed in the rabbits were enough to produce antibodies against Cryptosporidium, the standard direct immunofluorescence assay permitted the detection of five oocysts in 20% of the samples, and no cross-reaction with Giardia duodenalis cysts occurred.
Petersen, Heidi Huus; Enemark, Heidi L.; Olsen, Annette
The potential for transport of viable Cryptosporidium parvum oocysts through soil to land drains and groundwater was studied using simulated rainfall and intact soil columns which were applied raw slurry or separated liquid slurry. Following irrigation and weekly samplings over a four week period.......63) with the total number of oocysts found. Destructively sampling of the soil columns showed that type of slurry and irrigation played a role in the vertical distribution of oocysts, with more oocysts recovered from soil columns added liquid slurry irrespectively of irrigation status. Further studies are needed...
Lalancette, Cindy; Di Giovanni, George D; Prévost, Michèle
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.
Bonadonna, Lucia; Briancesco, Rossella; Ottaviani, Massimo; Veschetti, Enrico
Contaminated water are recognized as a potential source of Cryptosporidium oocysts. In Italy studies on the environmental spread of the parasite are still scarce even if a high frequency of serological responses could suggest high rates of endemic Cryptosporidium infections. In this investigation the occurrence of the parasite in water samples from a municipal wastewater treatment plant was monitored with the aim to evaluate possible correlation between oocysts and bacterial indicators, pathogens and physico-chemical parameters using a multivariate analysis. Data collected showed high numbers of oocysts, with a high percentage of positive samples and a significant correlation with pH, redox potential and total organic carbon. No correlation was found between Cryptosporidium concentration and that of the other microrganisms. The results, through a probing statistical analysis, show that none of the selected microrganisms was a reliable predictor of the presence of the parasite.
Schets, Franciska M; van den Berg, Harold H J L; de Roda Husman, Ana Maria
The intestinal parasites Cryptosporidium and Giardia are transmitted by water and food and cause human gastroenteritis. Filter-feeding bivalve mollusks, such as oysters and mussels, filter large volumes of water and thus concentrate such pathogens, which makes these bivalves potential vectors of disease. To assess the risk of infection from consumption of contaminated bivalves, parasite numbers and parasite recovery data are required. A modified immunomagnetic separation (IMS) procedure was used to determine Cryptosporidium oocyst and Giardia cyst numbers in individually homogenized oysters (Crassostrea gigas) and mussels (Mytilus edulis). About 12% of the commercial bivalves were positive, with low (oo)cyst numbers per specimen. The recovery efficiency of the IMS procedure was systematically evaluated. Experiments included seeding of homogenized bivalves and whole animals with 100 to 1,000 (oo)cysts. Both seeding procedures yielded highly variable recovery rates. Median Cryptosporidium recoveries were 7.9 to 21% in oysters and 62% in mussels. Median Giardia recoveries were 10 to 25% in oysters and 110% in mussels. Giardia recovery was significantly higher than Cryptosporidium recovery. (Oo)cysts were less efficiently recovered from seeded whole animals than from seeded homogenates, with median Cryptosporidium recoveries of 5.3% in oysters and 45% in mussels and median Giardia recoveries of 4.0% in oysters and 82% in mussels. Both bivalve homogenate seeding and whole animal seeding yielded higher (oo)cyst recovery in mussels than in oysters, likely because of the presence of less shellfish tissue in IMS when analyzing the smaller mussels compared with the larger oysters, resulting in more efficient (oo)cyst extraction. The data generated in this study may be used in the quantitative assessment of the risk of infection with Cryptosporidium or Giardia associated with the consumption of raw bivalve mollusks. This information may be used for making risk management
Amorós, Inmaculada; Alonso, José L; Cuesta, Gonzalo
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.
R. G. Mohammad
Full Text Available The study included examination of 110 water samples from well distributed in Mosul city and few towns and villages around it from May 2009 to March 2010 for detection of Cryptosporidium spp oocysts and Giardia spp cysts in well water. The results revealed that the prevalence of Cryptosporidium oocysts was 16.36% while the prevalence of Giardia cysts was 12.72%. The percentages of prevalence with Cryptosporidium and Giardia were in high rate in Bartilla and some villages around it 20% for Cryptosporidium and 17.14% for Giardia, the low rates were in Mosul city 10% for both protozoa. The highest prevalence rate of Cryptosporidium was in March 38.46% and the lowest was in November and July 0%. The highest prevalence rate of Giardia was in October 23.53% and the lowest rate in July 0%. This first study shows the presence of Cryptosporidium oocysts and Giardia cysts in well water (Ground water in Nineveh governorate.
Graczyk, Thaddeus K.; Fayer, Ronald; Cranfield, Michael R.; Conn, David Bruce
Asian freshwater clams, Corbicula fluminea, exposed for 24 h to 38 liters of water contaminated with infectious Cryptosporidium parvum oocysts (1.00 × 106 oocysts/liter; approximately 1.9 × 105 oocysts/clam) were examined (hemolymph, gills, gastrointestinal [GI] tract, and feces) on days 1, 2, 3, 7, and 14 postexposure (PE). No oocysts were detected in the water 24 h after the contamination event. The percentage of oocyst-containing clams varied from 20 to 100%, depending on the type of tissue examined and the technique used—acid-fast stain (AFS) or immunofluorescent antibody (IFA). The oocysts were found in clam tissues and feces on days 1 through 14 PE; the oocysts extracted from the tissues on day 7 PE were infectious for neonatal BALB/c mice. Overall, the highest number of positive samples was obtained when gills and GI tracts were processed with IFA (prevalence, 97.5%). A comparison of the relative oocyst numbers indicated that overall, 58.3% of the oocysts were found in clam tissues and 41.7% were found in feces when IFA was used; when AFS was used, the values were 51.9 and 48.1%, respectively. Clam-released oocysts were always surrounded by feces; no free oocysts or oocysts disassociated from fecal matter were observed. The results indicate that these benthic freshwater clams are capable of recovery and sedimentation of waterborne C. parvum oocysts. To optimize the detection of C. parvum oocysts in C. fluminea tissue, it is recommended that gill and GI tract samples be screened with IFA (such as that in the commercially available MERIFLUOR test kit). PMID:9464376
Méndez-Hermida, F; Castro-Hermida, J A; Ares-Mazás, E; Kehoe, S C; McGuigan, K G
The results of batch-process solar disinfection (SODIS) of Cryptosporidium parvum oocysts in water are reported. Oocyst suspensions were exposed to simulated sunlight (830 W m(-2)) at 40 degrees C. Viability assays (4',6'-diamidino-2-phenylindole [DAPI]/propidium iodide and excystation) and infectivity tests (Swiss CD-1 suckling mice) were performed. SODIS exposures of 6 and 12 h reduced oocyst infectivity from 100% to 7.5% (standard deviation = 2.3) and 0% (standard deviation = 0.0), respectively.
Petersen, Heidi H; Enemark, Heidi L; Olsen, Annette; Amin, M G Mostofa; Dalsgaard, Anders
The potential for the transport of viable Cryptosporidium parvum oocysts through soil to land drains and groundwater was studied using simulated rainfall and intact soil columns which were applied raw slurry or separated liquid slurry. Following irrigation and weekly samplings over a 4-week period, C. parvum oocysts were detected from all soil columns regardless of slurry type and application method, although recovery rates were low (vertical distribution of oocysts, with more oocysts recovered from soil columns added liquid slurry irrespective of the irrigation status. Further studies are needed to determine the effectiveness of different slurry separation technologies to remove oocysts and other pathogens, as well as whether the application of separated liquid slurry to agricultural land may represent higher risks for groundwater contamination compared to application of raw slurry.
Neumayerová, H.; Koudela, Břetislav
Roč. 153, 3/4 (2008), s. 197-202 ISSN 0304-4017 Institutional research plan: CEZ:AV0Z60220518 Keywords : Cryptosporidium muris * oocysts * low and high temperatures * infectivity Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 2.039, year: 2008
Previous studies have shown a dose-dependent effect of gamma irradiation on Cryptosporidium parvum development in neonatal mice and newborn calves. In mice, C. parvum oocysts exposed to 200 Gy showed nearly complete inability to develop as measured by C. parvum-specific quantitative PCR of ileal ti...
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...
Luo, Xia; Jedlicka, Sabrina S; Jellison, Kristen L
This study investigated Cryptosporidium parvum oocyst deposition onto biofilms as a function of shear stress under laminar or turbulent flow. Annular rotating bioreactors were used to grow stabilized stream biofilms at shear stresses ranging from 0.038 to 0.46 Pa. These steady-state biofilms were then used to assess the impact of hydrodynamic conditions on C. parvum oocyst attachment. C. parvum deposition onto biofilms followed a pseudo-second-order model under both laminar (after a lag phase) and turbulent flows. The total number of oocysts attached to the biofilm at steady state decreased as the hydrodynamic wall shear stress increased. The oocyst deposition rate constant increased with shear stress but decreased at high shear, suggesting that increasing wall shear stress results in faster attachment of Cryptosporidium due to higher mass transport until the shear forces exceed a critical limit that prevents oocyst attachment. These data show that oocyst attachment in the short and long term are impacted differently by shear: higher shear (to a certain limit) may be associated with faster initial oocyst attachment, but lower shear is associated with greater numbers of oocysts attached at equilibrium. IMPORTANCE This research provides experimental evidence to demonstrate that shear stress plays a critical role in protozoan-pathogen transport and deposition in environmental waters. The data presented in this work expand scientific understanding of Cryptosporidium attachment and fate, which will further influence the development of timely and accurate sampling strategies, as well as advanced water treatment technologies, to target protozoan pathogens in surface waters that serve as municipal drinking water sources. Copyright © 2017 American Society for Microbiology.
Kar, Sirri; Gawlowska, Sandra; Daugschies, Arwid; Bangoura, Berit
Cryptosporidium parvum (C. parvum) is the causal agent of cryptosporidiosis in many animals, mainly cattle, and possesses a high zoonotic potential. It occurs worldwide and ubiquitously. Detection of C. parvum is mainly performed directly but purification of the oocysts is useful to increase sensitivity and to obtain oocyst material for further use. The study was designed to compare (a) three different direct diagnostic methods, namely modified Ziehl-Neelsen staining, carbol fuchsin staining and conventional PCR, and (b) three routine oocyst purification methods, in particular flotation with saturated sodium chloride solution, Sheather's sucrose solution and a Percoll(®) gradient. During comparison of purification methods, special regard was paid to the ability to separate morphologically intact oocysts from the morphologically degenerated fraction or viable from non-viable oocysts, respectively. (a) Most effective in C. parvum oocysts detection in calf faeces was PCR; carbol fuchsin and modified Ziehl-Neelsen stainings achieved comparable results. (b) Purification methods: Oocyst flotation using sodium chloride solution showed to be superior to Percoll(®) gradient centrifugation and sugar flotation in terms of purification quality, recovery efficacy (yield) and reduction of the proportion of degenerated or non-viable oocysts. Copyright © 2010 Elsevier B.V. All rights reserved.
Background: Although Cryptosporidium spp. infections in acquired immunodeficiency syndrome patients (AIDS) with chronic diarrhoea have been reported in several African countries, there is no information regarding cryptosporidial diarrhoea in Ghanaian AIDS patients. Objective: To investigate the occurrence of C.
Robertson, L J; Gjerde, B
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.
Rahmatullah Rind, A.J. Probert1 and M.I. Rind2
Full Text Available Sixty three faecal as well as blood samples from a group of 15 young Friesian calves under 2 months of age at Aber Farm Bangor, U.K. were collected on monthly basis and examined for the presence of Cryptosporidium oocysts and serum immunoglobulin G (IgG antibodies, Twelve (19.23 % were found positive with Cryptosporium species while in 5 (7.9 % faecal samples both Cryptosporidium and Eimeria were present but 46 (73.0 % samples were negative. In 9 out of 12 (75.0 % cases where Cryptosporidium ocysts were present, a positive IF AT was observed while in 4 out of 5 (80.0 % positives were seen in the presence of both Cryptosporium and Eimeria oocysts. In contrast only 6 out of 46 (13.1% cases, a positive IFAT was also seen when no oocysts were recorded. Oocysts fluoresced brightly with positive serum samples and only faintly or not at all with the negative samples or the conjugate alone.
Schets FM; Medema GJ; Boschman GD; LWL; Becton Dickinson Europe, Aalst, Belgie
Current detection techniques for Cryptosporidium oocysts and Giardia cysts in water samples combine filtration of large volumes of water, concentration by centrifugation and flotation and immunofluorescense microscopy. The techniques are extremely labour-intensive and inefficient. The various steps
Ramirez, Norma E; Wang, Ping; Lejeune, Jeff; Shipitalo, Martin J; Ward, Lucy A; Sreevatsan, Srinand; Dick, Warren A
Most waterborne outbreaks of cryptosporidiosis have been attributed to agricultural sources due to the high prevalence of Cryptosporidium oocysts in animal wastes and manure spreading on farmlands. No-till, an effective conservation practice, often results in soil having higher water infiltration and percolation rates than conventional tillage. We treated six undisturbed no-till and six tilled soil blocks (30 by 30 by 30 cm) with 1 L liquid dairy manure containing 10(5) C. parvum oocysts per milliliter to test the effect of tillage and rainfall on oocyst transport. The blocks were subjected to rainfall treatments consisting of 5 mm or 30 mm in 30 min. Leachate was collected from the base of the blocks in 35-mL increments using a 64-cell grid lysimeter. Even before any rain was applied, approximately 300 mL of water from the liquid manure (30% of that applied) was transported through the no-till soil, but none through the tilled blocks. After rain was applied, a greater number and percentage of first leachate samples from the no-till soil blocks compared to the tilled blocks tested positive for Cryptosporidium oocysts. In contrast to leachate, greater numbers of oocysts were recovered from the tilled soil, itself, than from the no-till soil. Although tillage was the most important factor affecting oocyst transport, rainfall timing and intensity were also important. To minimize transport of Cryptosporidium in no-till fields, manure should be applied at least 48 h before heavy rainfall is anticipated or methods of disrupting the direct linkage of surface soil to drains, via macropores, need to be used.
Full Text Available Infection by Cryptosporidium baileyi causes respiratory cryptosporidiosis in red grouse Lagopus lagopus scotica. First diagnosed in 2010, it has since been detected across half of moors managed for grouse shooting in northern England. We hypothesised that contaminated grouse faeces within communal trays visited by grouse containing grit coated with flubendazole, provided to control Trichostrongylus tenuis parasites of grouse, is a reservoir of infection. To establish the basis to this hypothesis, contents of 23 trays from a grouse moor were examined for Cryptosporidium oocysts. Contents were subjected to Immuno Magnetic Separation oocyst concentration techniques prior to examination by Immuno Fluorescence Antibody Test microscopy and molecular analysis on the 18S rRNA gene. Seven of 13 (54% grit trays known to be used by infected grouse were positive for Cryptosporidium by IMS-IFAT, compared to two of 10 (20% random background trays. Ten of the 13 (77% trays used by infected birds amplified positive for Cryptosporidium by Polymerase Chain Reaction and three of the 10 (30% random trays. All PCR amplified products sequenced matched with C. baileyi, with C. parvum also present in one tray. These data suggest that trays used to “worm” grouse may act as reservoirs of Cryptosporidium infection and their future design may need to be reconsidered to minimise contamination.
Baines, David; Giles, Michaela; Richardson, Michael
Infection by Cryptosporidium baileyi causes respiratory cryptosporidiosis in red grouse Lagopus lagopus scotica . First diagnosed in 2010, it has since been detected across half of moors managed for grouse shooting in northern England. We hypothesised that contaminated grouse faeces within communal trays visited by grouse containing grit coated with flubendazole, provided to control Trichostrongylus tenuis parasites of grouse, is a reservoir of infection. To establish the basis to this hypothesis, contents of 23 trays from a grouse moor were examined for Cryptosporidium oocysts. Contents were subjected to Immuno Magnetic Separation oocyst concentration techniques prior to examination by Immuno Fluorescence Antibody Test microscopy and molecular analysis on the 18S rRNA gene. Seven of 13 (54%) grit trays known to be used by infected grouse were positive for Cryptosporidium by IMS-IFAT, compared to two of 10 (20%) random background trays. Ten of the 13 (77%) trays used by infected birds amplified positive for Cryptosporidium by Polymerase Chain Reaction and three of the 10 (30%) random trays. All PCR amplified products sequenced matched with C. baileyi , with C. parvum also present in one tray . These data suggest that trays used to "worm" grouse may act as reservoirs of Cryptosporidium infection and their future design may need to be reconsidered to minimise contamination.
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...
Although cysts and oocysts can be found in natural waters from any source, the risks of contamination are much greater in surface waters than in ground waters. A number of waterborne outbreaks of cryptosporidiosis, from potable and recreational water have been recorded. In many cases these outbreaks have occurred ...
Balthazard-Accou , Ketty; Emmanuel , Evens; Diouf , Momar; Agnamey , Patrice
International audience; Contamination of natural aquatic ecosystems by Cryptosporidium is a major environmental and human health issue. In Haiti, environmental Cryptosporidium oocysts pollution has been well documented by previous studies conducted in several cities of the country. In groundwater from Les Cayes of Haiti, significant concentrations from 1 to 989 oocysts in 100 liters of filtered water were calculated. Results of these studies revealed high level of Cryptosporidium oocysts poll...
Dec 2, 2014 ... Cryptosporidium, a coccidian protozoan parasite, is an important causative agent of human and animal gastrointestinal illness globally (Huang et al., 2009). ..... person, animal-to-person, waterborne, food-borne, and possible airborne transmission. This need to be further investigated. The observation made ...
Pecková, R.; Stuart, P. D.; Sak, Bohumil; Květoňová, Dana; Kváč, Martin; Foitová, I.
Roč. 230, OCT 30 (2016), s. 1-5 ISSN 0304-4017 R&D Projects: GA ČR(CZ) GAP505/11/1163 Institutional support: RVO:60077344 Keywords : Cryptosporidium parvum * excystation methods * in vitro cultivation * sodium hypochlorite * tlypsin Subject RIV: EG - Zoology Impact factor: 2.356, year: 2016
Also, no enterobacteria was found in the HIV-seronegative patients. Conclusion: This study demonstrates the prevalence of Cryptosporidium sp. in both HIV/ AIDS and HIV-seronegative individuals in Ghana. However, there was no statistical association between cryptosporidiosis and HIV/AIDS (p>0.05). East African ...
Salleh, Fatmah Md; Moktar, Norhayati; Yasin, Azlin Mohd; Al-Mekhlafi, Hesham M; Anuar, Tengku Shahrul
To improve the stool concentration procedure, we modified different steps of the standard formalin-ether concentration technique and evaluated these modifications by examining stool samples collected in the field. Seven samples were found positive by the modified formalin-ether concentration technique (M-FECT). Therefore, the M-FECT procedure provides enhanced detection of Cryptosporidium oocysts. Copyright © 2014 Elsevier B.V. All rights reserved.
Jeane Kury Nobre Gomes
Full Text Available The goal of this research was to verify the occurrence of Cryptosporidium spp. oocysts in the water consumption and supply of pools used by Antillean manatee (Trichechus manatus in captivity. Six samples were collected from the pool's supply system (at the beach, water used for manatee consumption, mineral water used in the preparation of artificial milk formulas offered to orphan manatee calves, also used in permanent maintenance pools in the visitation area, and water utilized in the rehabilitation area, where calves of all ages were kept. Before the water samples were processed, each sample was submitted to a filtration process. The diagnosis of the parasite was obtained by Kinyoun technique and the positive samples were submitted to the Direct Immunoflorescence Test. The results showed the presence of Cryptosporidium spp. oocysts in 66.67% (4/6 of the samples from the water supply system and pool, with eight to 56 oocysts per liter. There was evidence that the water used in the maintenance of the Antillean manatees could be an important medium for hydric transmission of Cryptosporidium spp. and that it may be a predisposing factor to the occurrence of cryptosporidiosis in Antillean manatees in captivity.
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.
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...... in the leachates from soil columns to which Cryptosporidium positive slurry had been injected. Although recovery rates were low, regardless of slurry type, C. parvum oocysts were detected from all soil columns. Variations in the leachate patterns were recorded between soil columns added raw and liquid slurry...
Silvia Cristina Osaki
Full Text Available INTRODUCTION: The production of anti-Cryptosporidium polyclonal antibodies and its use in direct immunofluorescence assays to determine the presence of Cryptosporidium in water are described in the present work. METHODS: Two rabbits were immunized with soluble and particulate antigens from purified Cryptosporidium oocysts. The sera produced were prepared for immunoglobulin G extraction, which were then purified and conjugated with fluorescein isothiocyanate (FITC. Slides containing known amounts of oocysts were prepared to determine the sensitivity of the technique. To test the specificity, slides containing Giardia duodenalis cysts were prepared. RESULTS: The conjugate was successfully used in water samples experimentally contaminated with Cryptosporidium oocysts, and it was possible to detect up to five oocysts/spot, corresponding to contamination of 250 oocysts/mL. CONCLUSIONS: The three immunizations performed in the rabbits were enough to produce antibodies against Cryptosporidium, the standard direct immunofluorescence assay permitted the detection of five oocysts in 20% of the samples, and no cross-reaction with Giardia duodenalis cysts occurred.
Al-Sabi, Mohammad Nafi Solaiman; Gad, Jens; Klinting, Mette
Background: The two most common water borne pathogenic protozoa, Cryptosporidium and Giardia, cause diarrhea worldwide. Detecting these parasites in water samples depends on effective parasite recovery from the water matrix. The reported low recovery rates of the currently used filter methods...... 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...
Christophe J. G. Darnault
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
Lim, Y A L; Aahmad, R A
A survey of the river water frequently used by the Temuan Orang Asli (aborigine) indicated that 66.7% of the river water samples were Giardia cyst positive and 5.6% were Cryptosporidium oocyst positive. Although Giardia cysts were detected in samples from all the sites (e.g. upstream, midstream, and downstream), Cryptosporidium was only present in one river water sample taken from downstream from a village. The only sample of upstream water which contained Giardia cysts had a concentration of 0.7 cysts/l. All samples taken from midstream contained cysts with a mean concentration of 9.8 +/- 6.6 cysts/l (range = 1-20 cysts/l). Eighty-three point three percent of the samples collected from downstream contained cysts and 16.7% had oocysts. The average concentration of cysts was 12.9 +/- 16.4 cysts/l (range = 0-44 cysts/l), whereas the oocyst concentration was 0.4 oocysts/l. All river samples tested positive for the presence of E. coli, indicating fecal contamination. The results of this study imply that the river system is contaminated with fecal-oral transmitted parasites. The river water, used by the Orang Asli, is a probable route for Giardia and Cryptosporidium transmission in this community. Long term strategies, incorporating health education regarding personal hygiene, and provision of toilets and the importance of their proper usage, need to be embraced by this community in order to control the spread of these parasites.
Park, Y.; Hou, L.; Atwill, R.; Packman, A. I.; Harter, T.
Cryptosporidium is one of the most common enteric parasites of humans and domestic animals, and a number of outbreaks of Cryprosporidiosis, a diarrheal disease caused by Cryptosporidium have been reported worldwide. Natural porous media has been demonstrated to be an effective filter for removing Cryptosporidium parvum from contaminated water and the amount of Cryptosporidium filtered is known to be highly dependent on physical and chemical conditions of the porous media and the water. Cryptosporidium deposition in saturated porous media involves two main steps: approach and attachment. In contrast to the approach mechanisms, attachment processes have not been systematically described to predict a priori because theories that represent attachment behavior (colloid stability) such as DLVO are insufficient to explain experimental data. For this reason, attachment efficiency is calculated based on empirical data, typically experimental breakthrough curves in laboratory columns or field experiments. In this study, collision (attachment) efficiencies (α) of C. parvum oocyst were calculated to test the effect of chemical property changes on the association of oocysts with sand grains. The breakthrough curve data obtained from twelve column experiments and three models were employed to calculate single collector efficiency (η) and α. The first ten experiments were conducted by changing ionic strength and pH, and mixing with natural sediments under the same physical properties (same η). Our experiment results show that iron coating or clay/suspended solids mixture drastically enhanced oocyst deposition. The experiments also showed that increase in ionic strength and decrease in pH enhanced the attachment efficiency. However, the experiment with 100mM NaCl resulted in low attachment efficiency and the experiment with pH 8.5 showed similar attachment efficiency to the one at pH 7. Based on the results from two additional experiments with different flow velocities, it
Kuczynska, E; Shelton, D R
Eight concentration and purification methods were evaluated to determine percentages of recovery of Cryptosporidium parvum oocysts from calf feces. The NaCl flotation method generally resulted in the highest percentages of recovery. Based on the percentages of recovery, the amounts of fecal debris in the final oocyst preparations, the relatively short processing time (flotation method was chosen for further evaluation. Extraction efficiency was evaluated by using oocyst concentrations of 25, 50, 10(2), 10(3), 10(4), and 10(5) oocysts g of bovine feces-1. The percentages of recovery ranged from 10.8% (25 oocysts g-1) to 17.0% (10(4) oocysts g-1) (r2 = 0.996). A conservative estimate of the detection limit for bovine feces is ca. 30 oocysts g of feces-1. Percentages of recovery were determined for six different types of animal feces (cow, horse, pig, sheep, deer, and chicken feces) at a single oocyst concentration (10(4) oocysts g-1). The percentages of recovery were highest for bovine feces (17. 0%) and lowest for chicken feces (3.2%). Percentages of recovery were determined for bovine manure after 3 to 7 days of storage. The percentages of recovery ranged from 1.9 to 3.5% depending on the oocyst concentration, the time of storage, and the dispersing solution. The percentages of oocyst recovery from soils were evaluated by using different flotation solutions (NaCl, cold sucrose, ZnSO4), different dispersing solutions (Triton X-100, Tween 80, Tris plus Tween 80), different dispersion techniques (magnetic stirring, sonication, blending), and different dispersion times (5, 15, and 30 min). Twenty-five-gram soil samples were used to reduce the spatial variability. The highest percentages of recovery were obtained when we used 50 mM Tris-0.5% Tween 80 as the dispersing solution, dispersion for 15 min by stirring, and saturated NaCl as the flotation solution. The percentages of oocyst recovery from freshly spiked sandy loam, silty clay loam, and clay loam soils were ca
Quintero-Betancourt, W; Gennaccaro, A L; Scott, T M; Rose, J B
This study evaluates the occurrence of Cryptosporidium oocysts and Giardia cysts in reclaimed effluents if method 1623 with the Envirochek capsule filters (standard and high-volume [HV] filters) and a modified version of the Information Collection Rule method (ICR) with the polypropylene yarn-wound cartridge filter are used. The recovery efficiency of the analytical methods was evaluated with samples of reagent, tap, and reclaimed water by using flow cytometer-sorted spike suspensions. (Oo)cyst recovery efficiency determined filter performance and method reproducibility in the water matrix tested. Method 1623 with the Envirochek HV capsule filter generated significantly higher recovery rates than did the standard Envirochek filter and the modified ICR method. Notwithstanding, large variations in recovery rates (>80%) occurred with samples of reclaimed water, and none of the water quality parameters analyzed in the reclaimed effluents could explain such variability. The highest concentrations of indigenous oocysts were detected by method 1623 with the HV filter, which provided a sufficient number of oocysts for further confirmation of infectious potential. Confirmation of species and potential infectivity for all positive protozoan samples was made by using a nested PCR restriction fragment polymorphism assay and the focus detection method most-probable-number assay, respectively. The methodology and results described in the present investigation provide useful information for the establishment of pathogen numeric standards for reclaimed effluents used for unrestricted irrigation.
Liu, Y.; Meng, X.; Guo, Z.; Zhang, C.; Nguyen, T. H.; Hu, D.; Ji, J.; Yang, X.
Colloidal attachment on charge heterogeneous grains has significant environmental implications for transport of hazardous colloids, such as pathogens, in the aquifer, where iron, manganese, and aluminium oxide minerals are the major source of surface charge heterogeneity of the aquifer grains. A patchwise surface charge model is often used to describe the surface charge heterogeneity of the grains. In the patchwise model, the colloidal attachment efficiency is linearly correlated with the fraction of the favorable patches (θ=λ(θf - θu)+θu). However, our previous microfluidic study showed that the attachment efficiency of oocysts of Cryptosporidium parvum, a waterborne protozoan parasite, was not linear correlated with the fraction of the favorable patches (λ). In this study, we developed a pore scale model to simulate colloidal transport and attachment on charge heterogeneous grains. The flow field was simulated using the LBM method and colloidal transport and attachment were simulated using the Lagrange particle tracking method. The pore scale model was calibrated with experimental results of colloidal and oocyst transport in microfluidic devices and was then used to simulate oocyst transport in charge heterogeneous porous media under a variety of environmental relative conditions, i.e. the fraction of favorable patchwise, ionic strength, and pH. The results of the pore scale simulations were used to evaluate the effect of surface charge heterogeneity on upscaling of oocyst transport from pore to continuum scale and to develop an applicable correlation between colloidal attachment efficiency and the fraction of the favorable patches.
Petersen, Heidi Huus; Petersen, T. B.; Enemark, Heidi
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 and stream water. Glass jars (n = 60) containing 500 mL wastewater obtained from the inlet to the primary settling tanks from.......7% reduction) in the treated wastewater and stream water, respectively. In contrast, the turbidity was 55.3 ± 4.4 NTU and 46.2 ± 1.6 NTU in untreated wastewater and stream water, respectively. M. oleifera seeds are readily available in many tropical countries where the tree is common, and our results clearly...
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...... in the leachates from soil columns to which Cryptosporidium positive slurry had been injected. Although recovery rates were low, regardless of slurry type, C. parvum oocysts were detected from all soil columns. Variations in the leachate patterns were recorded between soil columns added raw and liquid slurry...
Alonso, José L; Amorós, Inmaculada; Guy, Rebecca A
Real-time PCR (qPCR) is a rapid tool to quantify pathogens in the aquatic environment; however, it quantifies all pathogens, including both viable and nonviable. Propidium monoazide (PMA) is a membrane-impairment dye that penetrates only membrane-damaged cells. Once inside the cell, PMA is covalently cross-linked to DNA through light photoactivation, and PCR amplification is strongly inhibited. The goal of this study was to evaluate PMA-qPCR assays for rapid quantification of viable and heat-treated Giardia cysts and Cryptosporidium oocysts in wastewater. We observed a reduction in detection of heat-treated Giardia duodenalis cysts of 83.2, 89.9, 98.2, or 97% with PMA-qPCR assays amplifying a 75 base-pair (bp) β-giardin target, 77-bp triosephosphate isomerase (tpi), 133-bp glutamate dehydrogenase (GDH), and 143-bp β-giardin gene target, respectively. Thus, the exclusion of dead cysts was more effective when qPCR assays that produced larger amplicons were used. The PMA treatment of Cryptosporidium oocysts plus/minus heat treatment abolished the fluorescent signal for dead oocysts with a PMA-qPCR assay amplifying a Cryptosporidium parvum (150-bp) oocyst wall protein (COWP) gene. The PMA-qPCR 143-bp β-giardin assay for Giardia and the PMA-qPCR 150-bp COWP assay for Cryptosporidium accurately quantified live oo(cysts), and failed to detect dead oo(cysts), when phosphate-buffered saline and tertiary effluent wastewater were spiked with concentrations of 10(3) or 10(2) dead oo(cysts), respectively. Therefore, these assays are suitable for the detection of viable parasites that are typically present in tertiary wastewater effluents at concentrations of <10(3) oo(cysts)/l and can provide rapid risk assessments of environmental water.
Paziewska-Harris, A.; Schoone, G.; Schallig, H. D. F. H.
Viability estimation of the highly resistant oocysts of Cryptosporidium remains a key issue for the monitoring and control of this pathogen. We present here a simple 'one tube' quantitative PCR (qPCR) protocol for viability estimation using a DNA extraction protocol which preferentially solubilizes
Garcia, L S; Bruckner, D A; Brewer, T C; Shimizu, R Y
Due to increasing numbers of patients with documented infections with Cryptosporidium and other coccidia, it is important for the physician and clinical laboratory to be aware of the appropriate diagnostic techniques necessary for organism recovery and identification. Although Cryptosporidium is found in the gastrointestinal tract, tissue biopsies may be insufficient for organism recovery; the examination of stool specimens is a noninvasive procedure and will provide better overall opportunities for organism recovery. Human clinical specimens were examined from 45 patients with confirmed cryptosporidiosis or suspected of having the infection. Tissue biopsy sections, fecal wet preparations, and permanent stained smears were examined. Stool specimens were submitted in 10% Formalin, 2.5% potassium dichromate, and polyvinyl alcohol and were examined for oocysts by using 15 different methods: phase-contrast and light microscopy; Sheather's sugar flotation; Formalin concentration techniques; 10% potassium hydroxide; Giemsa; trichrome; periodic acid-Schiff; modified periodic acid-Schiff; silver methenamine; acridine orange; auramine-rhodamine; Kinyoun acid-fast; Ziehl-Neelsen carbolfuchsin; and a modified acid-fast procedure. Each technique or combination of techniques was assessed by organism quantitation, organism morphology, and ease of visual recognition. Based on these comparative studies, the modified Ziehl-Neelsen carbolfuchsin stain on 10% Formalin-preserved stool is recommended for the recovery and identification of Cryptosporidium.
Carlos J. Garro
Full Text Available In order to determine the prevalence and risk factors for shedding of Cryptosporidium spp. in dairy calves, a cross-sectional study was carried out in the northeastern region of Buenos Aires Province, Argentina. Fecal samples from a total of 552 calves from 27 dairy herds were collected, along with a questionnaire about management factors. Cryptosporidium spp. oocysts were detected by light microscopy using Kinyoun staining. Putative risk factors were tested for association using generalized linear mixed models (GLMMs. Oocyst shedding calves were found in 67% (CI95% = 49–84 of herds (corresponding to a true herd prevalence of 98% and 16% (CI95% = 13–19 of calves (corresponding to a true calve prevalence of 8%. Within-herd prevalence ranged from 0 to 60%, with a median of 8%. Cryptosporidium spp. excretion was not associated with the type of liquid diet, gender, time the calf stayed with the dam after birth, use of antibiotics, blood presence in feces, and calving season. However, important highly significant risk factors of oocyst shedding of calves was an age of less or equal than 20 days (OR = 7.4; 95% CI95% = 3–16; P < 0.0001 and occurrence of diarrhea (OR = 5.5; 95% CI95% = 2–11; P < 0.0001. The observed association with young age strongly suggests an early exposure of neonatal calves to Cryptosporidium spp. oocysts in maternity pens and/or an age-related susceptibility. Association with diarrhea suggests that Cryptosporidium spp. is an important enteropathogen primarily responsible for the cause of the observed diarrheal syndrome. Results demonstrate that Cryptosporidium spp. infection is widespread in the study region. Monitoring and control of this parasitic protozoan infection in dairy herds is recommended.
Méndez-Hermida, Fernando; Ares-Mazás, Elvira; McGuigan, Kevin G; Boyle, Maria; Sichel, Cosima; Fernández-Ibáñez, Pilar
The results of a batch-process solar disinfection (SODIS) and solar photocatalytic disinfection (SPCDIS) on drinking water contaminated with Cryptosporidium are reported. Cryptosporidium parvum oocyst suspensions were exposed to natural sunlight in Southern Spain and the oocyst viability was evaluated using two vital dyes [4',6-diamidino-2-phenylindole (DAPI) and propidium iodide (PI)]. SODIS exposures (strong sunlight) of 8 and 12h reduced oocyst viability from 98% (+/-1.3%) to 11.7% (+/-0.9%) and 0.3% (+/-0.33%), respectively. SODIS reactors fitted with flexible plastic inserts coated with TiO2 powder (SPCDIS) were found to be more effective than those which were not. After 8 and 16 h of overcast and cloudy solar irradiance conditions, SPCDIS reduced oocyst viability from 98.3% (+/-0.3%) to 37.7% (+/-2.6%) and 11.7% (+/-0.7%), respectively, versus to that achieved using SODIS of 81.3% (+/-1.6%) and 36.0% (+/-1.0%), respectively. These results confirm that solar disinfection of drinking water can be an effective household intervention against Cryptosporidium contamination.
Petersen, Tobias B; Petersen, Heidi H.; Abaidoo, Robert C.
Protozoan parasites belonging to the genus Cryptosporidium are transmitted e.g. by food and water and may cause severe diarrhoea, dehydration, weight loss and malnutrition. Ingestion of 10 oocysts can lead to infection and pathogenic symptoms. Thus, to characterize Cryptosporidium spp. contaminat...
Deming, Clare; Greiner, Ellis; Uhl, Elizabeth W
Cryptosporidiosis is an emerging problem in reptile medicine and has been associated with a wasting syndrome in leopard geckos (Eublepharis macularius). This study determined the prevalence of infection in a breeding colony of leopard geckos to be 9.8%. Two groups of 20 geckos, one that was fecal positive for oocysts of Cryptosporidium sp., and one, whose individuals were fecal negative at the inception of the study, were followed for 2 mo. Fecal samples were tested for oocysts every 2 wk, body weights were measured, and a body condition score was assigned for each gecko. Selected geckos from these two groups were euthanized and necropsied. There were statistically significant differences (P geckos with a subclinical or carrier state of infection. These animals continued to be infected with Cryptosporidium sp. but gained weight and remained in good body condition. Only one gecko in the entire group of 40 was confirmed to be negative for oocysts or developmental stages by repeated fecal exams and histopathology. An additional 37 severely emaciated geckos from the breeding colony were euthanized, and all were positive for Cryptosporidium sp. on histopathologic examination of the gastrointestinal tract. The results of this study indicate that although some animals can recover from a clinical infection, if a gecko is severely wasted, it should be euthanized because of the poor prognosis and possible source of infection to other geckos.
Mason, B W; Chalmers, R M; Carnicer-Pont, D; Casemore, D P
An outbreak in the autumn of 2005 resulted in 218 confirmed cases of Cryptosporidium hominis. The attack rate (relative risk 4.1, 95%CI 2.8-9.1) was significantly higher in the population supplied by Cwellyn Water Treatment Works (WTW). A case-control study demonstrated a statistically significant association (odds ratio 6.1, 95% CI 1.8-23.8) between drinking unboiled tap water and C. hominis infection. The association remained significant in a logistic regression analysis, with an adjusted odds ratio of 1.30 (95 CI 1.05-1.61) per glass of unboiled tap water consumed per day. This evidence together with environmental and associated microbiological investigations, and the absence of effective treatment to remove Cryptosporidium oocysts at the WTW, led to the conclusion that the outbreak was waterborne. Oocyst counts in final treated water at the WTW and at different points in the distribution system were consistently very low, maximum count in continuous monitoring 0.08 oocysts per 10 litres. Data from continuous monitoring and the epidemic curve is consistent with the hypothesis that low numbers of oocysts of C hominis were present in treated water continuously during the outbreak and these were of sufficient infectivity to cause illness. All surface water derived water supplies present a potential risk to human health and appropriate control measures should be in place to minimise these risks.
Schets FM; Engels GB; During M; Roda Husman AM de; MGB
Cryptosporidium is one of the important causative agents of gastrointestinal illness in humans. Cryptosporidium infections are often waterborne and can be transmitted through drinking water or recreational water. Estimation of the risk of infection with Cryptosporidium after exposure to drinking
Abbassi, H; Wyers, M; Cabaret, J; Naciri, M
A simple semiquantitative microscopic slide flotation (MSF) method using modified Sheather's sugar solution (MSSS) is presented for the rapid detection and quantification of Cryptosporidium baileyi oocysts in the feces and mucosal and/or organ scrapings of chickens. Oocyst shedding was evaluated by examination of the surface of coverslips, and the average quantitative score (0-5) recorded for 10 microscopic fields (magnification x250) is reported. The equivalence between these scores and the actual number of oocysts counted per gram of feces was assessed (rs = 0.89; P < 0.001). The applicability of this method was tested by comparison of the kinetics of oocyst shedding in feces of inoculated chickens with those reported by other authors working under similar conditions. In organs the MSF method was compared to histology. Fewer false-negative results were obtained using MSF versus the histology method. The MSF method was particularly more efficient in tracheae with low levels of infection and in the lungs, regardless of the level of infection. The MSF method was also very efficient in detecting oocysts in air sacs from chickens with aerosacculitis. It provides a specific and sufficiently sensitive, simple, rapid, reliable, and low-cost means of diagnosing C. baileyi in the feces and organs of chickens. This method can be used in the routine diagnosis of cryptosporidia in chickens, and it could be extended to other avian species and used in epidemiology studies to evaluate the prevalence of cryptosporidiosis in fowl.
Mohanram, Arvind; Ray, Chittaranjan; Harvey, Ronald W.; Metge, David W.; Ryan, Joseph N.; Chorover, Jon; Eberl, D.D.
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
Mohanram, A.; Ray, C.; Harvey, R.W.; Metge, D.W.; Ryan, J.N.; Chorover, J.; Eberl, D.D.
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
Hijnen, W.A.M.; Beerendonk, E.F.; Medema, Gerriet Jan
UV disinfection technology is of growing interest in the water industry since it was demonstrated that UV radiation is very effective against (oo)cysts of Cryptosporidium and Giardia, two pathogenic micro-organisms of major importance for the safety of drinking water. Quantitative Microbial Risk
Schets FM; Engels GB; During M; de Roda Husman AM; MGB
Cryptosporidium is een van de belangrijkste veroorzakers van gastro-enteritis bij de mens. Cryptosporidium-infecties worden vaak via water overgedragen, dit kan zowel drinkwater als recreatiewater zijn. Bij schatting van de kans op infectie met Cryptosporidium na blootstelling aan drinkwater is
Di Giovanni, George D.; Rochelle, Paul A.
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
Johnson, Anne M; Giovanni, George D Di; Rochelle, Paul A
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.
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.
Lu, Ping; Amburgey, James E; Hill, Vincent R; Murphy, Jennifer L; Schneeberger, Chandra L; Arrowood, Michael J; Yuan, Tao
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.
Garcia, L S; Bruckner, D A; Brewer, T C; Shimizu, R Y
Due to increasing numbers of patients with documented infections with Cryptosporidium and other coccidia, it is important for the physician and clinical laboratory to be aware of the appropriate diagnostic techniques necessary for organism recovery and identification. Although Cryptosporidium is found in the gastrointestinal tract, tissue biopsies may be insufficient for organism recovery; the examination of stool specimens is a noninvasive procedure and will provide better overall opportunit...
Full Text Available Giardia duodenalis, Cryptosporidium spp. and Toxoplasma gondii are protozoan parasites that have been highlighted as emerging foodborne pathogens by the Food and Agriculture Organization of the United Nations and the World Health Organization. According to the European Food Safety Authority, 4786 foodborne and waterborne outbreaks were reported in Europe in 2016, of which 0.4% were attributed to parasites including Cryptosporidium, Giardia and Trichinella. Until 2016, no standardized methods were available to detect Giardia, Cryptosporidium and Toxoplasma (oocysts in food. Therefore, no regulation exists regarding these biohazards. Nevertheless, considering their low infective dose, ingestion of foodstuffs contaminated by low quantities of these three parasites can lead to human infection. To evaluate the risk of protozoan parasites in food, efforts must be made towards exposure assessment to estimate the contamination along the food chain, from raw products to consumers. This requires determining: (i the occurrence of infective protozoan (oocysts in foods, and (ii the efficacy of control measures to eliminate this contamination. In order to conduct such assessments, methods for identification of viable (i.e. live and infective parasites are required. This review describes the methods currently available to evaluate infectivity and viability of G. duodenalis cysts, Cryptosporidium spp. and T. gondii oocysts, and their potential for application in exposure assessment to determine the presence of the infective protozoa and/or to characterize the efficacy of control measures. Advantages and limits of each method are highlighted and an analytical strategy is proposed to assess exposure to these protozoa.
An optimized cell culture-immunofluorescence (IFA) procedure, using the HCT-8 cell line, was evaluated in 'blind' trials to determine the sensitivity and reproducibility for measuring infectivity of flow cytometry prepared inocula of C. parvum oocysts. In separate trials, suspens...
Full Text Available per sample and the number of positive samples. Almost 50% of sewage samples studied contained Giardia cysts and 30% contained both Giardia cysts and Cryptosporium oocysts. Treatment of sewage resulted in a reduction in the percentage of samples...
Kváč, Martin; Květoňová, Dana; Salát, Jiří; Ditrich, Oleg
Roč. 100, č. 2 (2007), s. 213-217 ISSN 0932-0113 R&D Projects: GA ČR GA524/05/0992 Institutional research plan: CEZ:AV0Z60220518 Keywords : Cryptosporidium andersoni * viability * infectivity * long-term storage Subject RIV: EG - Zoology Impact factor: 1.512, year: 2007
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...
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 ...
Liu, Yuanyuan; Zhang, Changyong; Hu, Dehong; Kuhlenschmidt, Mark S.; Kuhlenschmidt, Theresa B.; Mylon, Steven E.; Kong, Rong; Bhargava, Rohit; Nguyen, Thanh H.
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.
Al-Sabi, Mohammad Nafi Solaiman; Gad, J.; Klinting, M.
Background: Sonication of fluids has been widely used for cell lyses and decontamination of water and food due to its three major physical and chemical effects: cavitation, implosion of water bubbles with subsequent increase in substrate temperature and liberation of free radicals such as O...... power and time durations on two of the most common waterborne protozoa Cryptosporidium and Giardia, and examined its effects on parasite characteristics and survival rate using immunofluorescence dyes; DAPI (4’,6-diamidino-2-phenylindol) staining/PI (propidium iodide), and analyzed by flow cytometry...... determine the effects of ultrasound on the parasite, including the sonication power of ultrasound as well as substrate temperature. Conclusions: Ultrasound is harmful for waterborne protozoa even when momentarily applied. However, a mode of operation may exist in which ultrasound can be used for collection...
Majewska, Anna C; Graczyk, Thaddeus K; Słodkowicz-Kowalska, Anna; Tamang, Leena; Jedrzejewski, Szymon; Zduniak, Piotr; Solarczyk, Piotr; Nowosad, Andrzej; Nowosad, Piotr
As Cryptosporidium parvum and Giardia lamblia can be disseminated in the environment by avian hosts, a total of 499 fecal dropping from 308 free-ranging, 90 captive, and 101 domestic birds were tested by conventional, immunological, and molecular techniques for these human enteropathogens. Twenty-six (5.2%) tested positive for G. lamblia cysts and 19 (3.8%) for C. parvum oocysts. A bird total of 23 (7.5%) free-ranging, two (2.2%) captive, and one (0.1%) domestic tested positive for cysts, whereas 18 (5.8%) free-ranging, one (1.1%) captive, and zero livestock birds tested positive for oocysts. G. lamblia cysts and C. parvum oocysts were found significantly more frequently in fecal droppings of free-ranging aquatic birds than in birds not normally associated with water. No specimen tested positive for both pathogens simultaneously. Aquatic birds represent an important epidemiologic link in water-associated transmission cycles of Cryptosporidium and Giardia and play a significant role in environmental contamination of aquatic habitats with these anthropozoonotic pathogens.
The effect of UV exposure on Toxoplasma gondii oocysts has not been completely defined for use in water disinfection. This study evaluated UV irradiated oocysts by three assays: a SCID mouse bioassay, an in vitro T. gondii oocyst plaque assay (TOP-assay), and a quantitative reve...
Of nearly 25 named species and numerous genotypes of Cryptosporidium, two are of special importance relative to human health and food safety: Cryptosporidium hominis and Cryptosporidium parvum, the former with a predilection for humans and the latter a promiscuous species. Genetic tools have been es...
Results show that both Cryptosporidium and Giardia attach to biofilms in large numbers (100-1000 oo/cysts) in as little as 72 hours. Pulsed light successfully inactivated all test species (Listeria, Salmonella, Bacillus, Escherichia) in planktonic and biofilm form with an increase in inactivation for every increase in UV dose.
Siqueira-Castro, Isabel Cristina Vidal; Greinert-Goulart, Juliane Araújo; Bonatti, Tais Rondello; Yamashiro, Sandra; Franco, Regina Maura Bueno
Ciliated protozoa are important components of the microbial food web in various habitats, especially aquatic environments. These organisms are useful bioindicators for both environmental quality assessment and the wastewater purification process. The pathogenic parasitic protozoan species Giardia and Cryptosporidium represent a significant concern for human health, being responsible for numerous disease outbreaks worldwide. The predation of cysts and oocysts in 15 ciliate species from water and sewage samples collected in Campinas, São Paulo, Brazil were verified under laboratory conditions. The ciliated protozoan species were selected based on their mode of nutrition, and only bacterivorous and suspension-feeders were considered for the experiments. The species Blepharisma sinuosum, Euplotes aediculatus, Sterkiella cavicola, Oxytricha granulifera, Vorticella infusionum, Spirostomum minus, and Stentor coeruleus ingested cysts and oocysts, the resistance forms of Giardia spp. and Cryptosporidium spp., respectively. This is the first time that the ingestion of Giardia cysts by ciliated protozoa has been reported. These findings may contribute to a better understanding of the biological removal of these pathogens from aquatic environments.
Comparison of propidium monoazide-quantitative PCR and reverse transcription quantitative PCR for viability detection of fresh Cryptosporidium oocysts following disinfection and after long-term storage in water samples
Purified oocysts of Cryptosporidium parvum were used to evaluate applicability of two quantitative PCR (qPCR) viability detection methods in raw surface water and disinfection treated water. Propidium monoazide-qPCR targeting hsp70 gene was compared to reverse transcription (RT)-...
Occurrence of Giardia cysts and Cryptosporidium oocysts in sewage and their reduction in treatment process; Gesuichu no Giardia shisuto oyobi Cryptosporidium oshisuto noudo to gesui syori ni okeru jyokyosei
Hashimoto, A.; Hirata, T. [Azabu University, Kanagawa (Japan); Magara, Y. [National Institute of Public Health, Tokyo (Japan); Ogaki, S. [The University of Tokyo, Tokyo (Japan)
Recently, the water system infection owing to protozoan, virus and so on which is different from the conventional infection like represented by bacteria, is reported. There are few examples for the infection conditional study and epidemiological investigation of these protozoan and virus in Japan. Authors carried out a field survey at nine sewage treatment plants, in order to investigate the occurrence of Giardia cysts and Cryptosporidium oocysts in raw sewage and the removal in sewage treatment processes. As a result, in all plants, Giardia cysts were detected in raw sewage and effluent of primary sedimentation. Cryptosporidium oocysts were detected from only one raw sewage sample. Indicator bacteria counts tested, C.perfringens spores, coliforms and E.coli, were statistically correlated with Giardia cyst densities and the most significant relationship was observed between C.perfringens spore counts and Giardia cyst densities. It was found that turbidity was the most appropriate indicator for Giardia cyst removal in activated sludge treatment process. 18 refs., 6 figs., 2 tabs.
The Iowa strain of Cryptosporidium parvum will not propagate in immunocompetent mice, but will successfully infect genetically immunocompromised Nude or SCID mice as well as immunocompetent mice which have been immunosuppressed with glucocorticoids. Using dexamethasone - tetracy...
...) and viruses under the Surface Water Treatment Rule (SWTR). Minimum CT requirements include relatively large safety factors to account for possible deviations from actual disinfection efficiencies achieved in full-scale contactors...
U.S.EPA Methods 1622 and 1623 are used for the detection of waterborne Cryptosporium. These methods consist of filtration, elution, purificaiton by immunomagnetic separation (IMS), and microscopic analysis for oocysts stained by a fluorescent monoclonal antibody and counter stai...
Schets FM; Medema GJ; Boschman GD; LWL; Becton Dickinson Europe; Aalst; Belgie
De huidige methode die in de Verenigde Staten, Groot- Brittannie en Nederland wordt gebruikt voor de detectie van Cryptosporidium en Giardia in water bestaat uit een combinatie van filtratie, concentratie door centrifugeren en flotatie en immuunfluorescentie microscopie. De methode is zeer
Occurrence of Giardia cysts and Cryptosporidium oocysts in activated sludge samples in Campinas, SP, Brazil Ocorrência de cistos de Giardia e oocistos de Cryptosporidium em amostras de lodo ativado em Campinas, SP, Brasil
Luciana Urbano Santos
Full Text Available Giardia and Cryptosporidium have caused several outbreaks of gastroenteritis in humans associated with drinking water. Contaminated sewage effluents are recognized as a potential source of waterborne protozoa. Due to the lack of studies about the occurrence of these parasites in sewage samples in Brazil, we compared the efficiency of two procedures for concentrating cysts and oocysts in activated sludge samples of one sewage treatment plant. For this, the samples were submitted to i concentration by the ether clarification procedure (ECP and to ii purification by sucrose flotation method (SFM and aliquots of the pellets were examined by immunofluorescence. Giardia cysts were present in all samples (100.0%; n = 8 when using ECP and kit 1 reagents, while kit 2 resulted in six positive samples (85.7%; n = 7. As for SFM, cysts were detected in 75.0% and 100.0% of these samples (for kit 1 and 2, respectively. Regarding Cryptosporidium, two samples (25.0%; kit 1 and 28.5% for kit 2 were detected positive by using ECP, while for SFM, only one sample (examined by kit 1 was positive (12.5%. The results of the control trial revealed Giardia and Cryptosporidium recovery efficiency rates for ECP of 54.5% and 9.6%, while SFM was 10.5% and 3.2%, respectively. Considering the high concentration detected, a previous evaluation of the activated sludge before its application in agriculture is recommended and with some improvement, ECP would be an appropriate simple technique for protozoa detection in sewage samples.Giardia e Cryptosporidium causaram vários surtos epidêmicos de gastroenterite, associados à água potável. Efluentes de esgoto contaminados foram incriminados como uma fonte potencial de cistos e oocistos. Uma investigação foi conduzida para verificar a presença de cistos de Giardia e oocistos de Cryptosporidium em amostras de lodo ativado de uma Estação de Tratamento de Esgoto. Para isto as amostras foram submetidas: i a concentração pelo
Petersen, Heidi Huus; Woolsey, Ian David; Dalsgaard, Anders
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 ...... water free of C. parvum oocysts as well as other protozoan parasites.......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...... in the untreated wastewater. Furthermore, the number of oocysts L-1was significantly (pwater, which was significantly (p
Le Goff, L; Hubert, B; Favennec, L; Villena, I; Ballet, J J; Agoulon, A; Orange, N; Gargala, G
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
Full Text Available Maturation ponds are used in rural and regional areas in Australia to remove the microbial loads of sewage wastewater, however, they have not been studied intensively until present. Using a combination of culture-based methods and quantitative real-time PCR, we assessed microbial removal rates in maturation ponds at four waste stabilization ponds (WSP with (n = 1 and without (n = 3 baffles in rural and remote communities in Australia. Concentrations of total coliforms, E. coli, enterococci, Campylobacter spp., Salmonella spp., F+ RNA coliphage, adenovirus, Cryptosporidium spp. and Giardia (oo cysts in maturation ponds were measured at the inlet and outlet. Only the baffled pond demonstrated a significant removal of most of the pathogens tested and therefore was subjected to further study by analyzing E. coli and enterococci concentrations at six points along the baffles over five sampling rounds. Using culture-based methods, we found a decrease in the number of E. coli and enterococci from the initial values of 100,000 CFU per 100 mL in the inlet samples to approximately 1000 CFU per 100 mL in the outlet samples for both bacterial groups. Giardia cysts removal was relatively higher than fecal indicators reduction possibly due to sedimentation.
Detecção de oocistos de Cryptosporidium spp. e cistos de Giardia spp. em amostras de esgoto bruto ou tratado: avaliação crítica dos métodos Detection of Cryptosporidium spp. oocysts and Giardia spp. cysts in raw and effluent wastewater: critical evaluation of methods
Luciana Urbano dos Santos
Full Text Available Neste trabalho, avaliou-se a eficiência dos métodos centrífugo-concentração e filtração em membrana, na detecção de oocistos de Cryptosporidium spp. e cistos de Giardia spp. em amostras de esgoto bruto e tratado, provenientes de um sistema de lodos ativados (estação de tratamento de esgoto, Samambaia, Campinas, em São Paulo. As amostras foram coletadas quinzenalmente por dois anos: 53 amostras de esgoto bruto (AFL, 53 de efluente tratado sem desinfecção por luz ultravioleta (EFL e 38 de efluente tratado e desinfetado por luz ultravioleta (EFL+UV. Cistos de Giardia spp. foram encontrados em 90,5% das amostras AFL; em 96,2%, de EFL; e em 94,7%, de EFL+UV. Oocistos de Cryptosporidium spp. foram detectados em 6,4% das amostras AFL e em 2,6 % de EFL+UV. Ambos os métodos mostraram-se eficientes na detecção destes protozoários em todos os tipos de amostras, além de apresentarem baixo custo por análise.In this study, the efficiency of centrifuge-concentration and membrane filtrated methods was evaluated in the detection of Cryptosporidium spp. oocysts and Giardia spp. cysts in raw or treated wastewater samples, from activated sludge systems (ETE - Samambaia, Campinas, in São Paulo. The samples were collected once a fortnight for two years: 53 samples of influent (AFL, 53 samples of treated effluent without ultraviolet disinfection (EFL, and 38 samples of treated effluent with ultraviolet disinfection (EFL+UV. Giardia spp. cysts were found in 90.5% of the AFL samples; in 96.2% of the samples, EFL; and in 94.7%, EFL+UV. Cryptosporidium spp. oocysts were detected in 6.4% of AFL samples and 2.6% of EFL+UV. Both methods showed efficiency when detecting protozoa in all types of samples, besides having low costs by analysis.
Tanaka, Y.; Takahashi, K. [Fuji Electric Co. Ltd., Tokyo (Japan); Motoyama, N. [Fuji Electric Corporate Research and Development, Ltd., Kanagawa (Japan)
Measures against Cryptosporidium parvum (C. parvum) in the waterworks are discussed. C. parvum is a pathogenic protozoan, and exists in the form of oocyst protected by a hard shell. It does not multiply in water or food, but does in human intestines and causes violent diarrhea and bellyache. A grave concern was created when many people were infected with the protozoan via tap water in Japan and the United States. Under such circumstances, ozone is used in an experiment to inactivate C. parvum. It is found that the C. parvum oocyst inactivation effect is evaluated by using a Ct value (disinfectant concentration Cmg/Ltimescontact time in minute) and that ozone treatment inactivates 90-99% of the protozoan. When various advanced water treatment technologies are being introduced for the purpose of serving safe and tasty water, the outcome of this study conveniently offers an ozone treatment method that will additionally inactivate pathogenic protozoa. Studies will be continued to elucidate the effects of factors of ozone treatment and water quality for the completion of an ideal disinfection process. Reference is made to an example of disinfection work implemented at a water purification plant of Milwaukie City, United States. 9 refs., 6 figs., 4 tabs.
Schrijven JF; Bruin HAM de; Engels GB; Leenen EJTM; MGB
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)
Ortega, Ynes R; Torres, Maria P; Tatum, Jessica M
Foodborne parasites are characterized as being highly resistant to sanitizers used by the food industry. In 2009, a study reported the effectiveness of levulinic acid in combination with sodium dodecyl sulfate (SDS) in killing foodborne bacteria. Because of their innocuous properties, we studied the effects of levulinic acid and SDS at various concentrations appropriate for use in foods, on the viability of Cryptosporidium parvum and Encephalitozoon intestinalis. The viability of Cryptosporidium and E. intestinalis was determined by in vitro cultivation using the HCT-8 and RK-13 cell lines, respectively. Two Escherichia coli O157:H7 isolates were also used in the present study: strain 932 (a human isolate from a 1992 Oregon meat outbreak) and strain E 0018 (isolated from calf feces). Different concentrations and combinations of levulinic acid and SDS were tested for their ability to reduce infectivity of C. parvum oocysts (10(5)), E. intestinalis spores (10(6)), and E. coli O157:H7 (10(7)/ml) when in suspension. Microsporidian spores were treated for 30 and 60 min at 20 ± 2°C. None of the combinations of levulinic acid and SDS were effective at inactivating the spores or oocysts. When Cryptosporidium oocysts were treated with higher concentrations (3% levulinic acid-2% SDS and 2% levulinic acid-1% SDS) for 30, 60, and 120 min, viability was unaffected. E. coli O157:H7, used as a control, was highly sensitive to the various concentrations and exposure times tested. SDS and levulinic acid alone had very limited effect on E. coli O157:H7 viability, but in combination they were highly effective at 30 and 60 min of incubation. In conclusion, Cryptosporidium and microsporidia are not inactivated when treated for various periods of time with 2% levulinic acid-1% SDS or 3% levulinic acid-2% SDS at 20°C, suggesting that this novel sanitizer cannot be used to eliminate parasitic contaminants in foods.
Khaldi, S; Ratajczak, M; Gargala, G; Fournier, M; Berthe, T; Favennec, L; Dupont, J P
Groundwater from karst aquifers is an important source of drinking water worldwide. Outbreaks of cryptosporidiosis linked to surface water and treated public water are regularly reported. Cryptosporidium oocysts are resistant to conventional drinking water disinfectants and are a major concern for the water industry. Here, we examined conditions associated with oocyst transport along a karstic hydrosystem, and the impact of intensive exploitation on Cryptosporidium oocyst contamination of the water supply. We studied a well-characterized karstic hydrosystem composed of a sinkhole, a spring and a wellbore. Thirty-six surface water and groundwater samples were analyzed for suspended particulate matter, turbidity, electrical conductivity, and Cryptosporidium and Giardia (oo)cyst concentrations. (Oo)cysts were identified and counted by means of solid-phase cytometry (ChemScan RDI(®)), a highly sensitive method. Cryptosporidium oocysts were detected in 78% of both surface water and groundwater samples, while Giardia cysts were found in respectively 22% and 8% of surface water and groundwater samples. Mean Cryptosporidium oocyst concentrations were 29, 13 and 4/100 L at the sinkhole, spring and wellbore, respectively. Cryptosporidium oocysts were transported from the sinkhole to the spring and the wellbore, with respective release rates of 45% and 14%, suggesting that oocysts are subject to storage and remobilization in karst conduits. Principal components analysis showed that Cryptosporidium oocyst concentrations depended on variations in hydrological forcing factors. All water samples collected during intensive exploitation contained oocysts. Control of Cryptosporidium oocyst contamination during intensive exploitation is therefore necessary to ensure drinking water quality. Copyright © 2011. Published by Elsevier Ltd.
Petersen, T. B.; Petersen, H. H.; Abaidoo, R. C.
Protozoan parasites belonging to the genus Cryptosporidium are transmitted e.g. by food and water and may cause severe diarrhoea, dehydration, weight loss and malnutrition. Ingestion of 10 oocysts can lead to infection and pathogenic symptoms. Thus, to characterize Cryptosporidium spp. contaminat...
A. Dellarupe; J.M. Unzaga; G. Moré; M. Kienast; A. Larsen; C. Stiebel; M. Rambeaud; M.C. Venturini
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 varani...
Liu, Xuehan; He, Tingmei; Zhong, Zhijun; Zhang, Hemin; Wang, Rongjun; Dong, Haiju; Wang, Chengdong; Li, Desheng; Deng, Jiabo; Peng, Guangneng; Zhang, Longxian
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. © 2013 Elsevier Ireland Ltd. All rights reserved.
Deng, M Q; Cliver, D O
Cryptosporidium parvum is a protozoan parasite capable of causing massive waterborne outbreaks. This study was conducted to model the transfer of C. parvum oocysts from contaminated water via food contact surfaces into yogurt and ice-cream, as well as to examine oocyst survival. Propidium iodide staining, combined with a direct immunofluorescence assay, was used for oocyst viability determination. Oocysts were recovered from milk products by a sucrose flotation-based procedure, with average recoveries of 82.3, 60.7, and 62.5% from low (1%) fat milk, 9% fat ice-cream, and 98% fat-free yogurt, respectively. Oocysts were also recovered, by rinsing with tap water, from stainless steel surfaces inoculated with oocyst suspension, with average recoveries of 93.1% when the surface was still wet and 69.0% after the surface had air-dried at room temperature. Viability of oocysts on the surface was significantly affected by desiccation; 5% of the oocysts remained viable after 4 h of air-drying at room temperature, while the proportion of viable oocysts was 81, 69, and 45% after air-drying for 10 min, 1 h, and 2 h, respectively. In contrast, oocyst viability only dropped from 82 to 75% after 30 min contact at room temperature with 5% bleach solution (equivalent to 0.26% NaOCl). Transfer of oocysts from milk and stainless steel surfaces into yogurt, and oocyst survival during the process were analyzed. Yogurt was made from pasteurized low fat milk and live yogurt starter by incubating at 37 degrees C for 48 h and then stored at 4 degrees C. Oocyst viability decreased from 83% (80%) to approximately 60% after 48 h at 37 degrees C and to approximately 58% following 8 days of storage, similar to oocyst survival in the controls using pasteurized milk without the addition of live yogurt. Oocyst survival in ice-cream was investigated by inoculating oocysts into ice-cream mix, and mixing and freezing in an ice-cream freezer, and hardening at -20 degrees C. Although approximately 20
Baishanbo, Asiya; Gargala, Gilles; Delaunay, Agnès; François, Arnaud; Ballet, Jean-Jacques; Favennec, Loïc
One-month-old dexamethasone-immunosuppressed Mongolian gerbils were challenged with 1 oocyst to 2 × 105 oocysts from two isolates genotyped as Cryptosporidium hominis and C. parvum (genotype 2), respectively. A similar dose-dependent gut infection was obtained, and the initial genotype maintained for 21 to 22 days. The data suggest that immunosuppressed gerbils provide a reliable rodent model of persistent C. hominis infection.
Enemark, Heidi L.; Ahrens, Peter; Juel, Cynthia Dawn
The genetic polymorphism among 271 Danish Cryptosporidium isolates of human and animal origin was studied by partial amplification and sequencing of the Cryptosporidium oocyst wall protein (COWP) gene, the 18S rDNA, and a microsatellite locus.dagger Furthermore, the microsatellite locus was studied...... (P Cryptosporidium isolates of human origin the anthroponotic subgenotype H1 was identified, in addition to the zoonotic subgenotypes C1, C2, and C3. Of 44 human samples, 56.8% were anthroponotic, whereas 40.9% were zoonotic genotypes. One human isolate...... 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...
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 ...
Vermeulen, Lucie C; Benders, Jorien; Medema, Gertjan; Hofstra, Nynke
Understanding the environmental pathways of Cryptosporidium is essential for effective management of human and animal cryptosporidiosis. In this paper we aim to quantify livestock Cryptosporidium spp. loads to land on a global scale using spatially explicit process-based modeling, and to explore the effect of manure storage and treatment on oocyst loads using scenario analysis. Our model GloWPa-Crypto L1 calculates a total global Cryptosporidium spp. load from livestock manure of 3.2 × 10 23 oocysts per year. Cattle, especially calves, are the largest contributors, followed by chickens and pigs. Spatial differences are linked to animal spatial distributions. North America, Europe, and Oceania together account for nearly a quarter of the total oocyst load, meaning that the developing world accounts for the largest share. GloWPa-Crypto L1 is most sensitive to oocyst excretion rates, due to large variation reported in literature. We compared the current situation to four alternative management scenarios. We find that although manure storage halves oocyst loads, manure treatment, especially of cattle manure and particularly at elevated temperatures, has a larger load reduction potential than manure storage (up to 4.6 log units). Regions with high reduction potential include India, Bangladesh, western Europe, China, several countries in Africa, and New Zealand.
Gómez-Couso, Hipólito; Fontán-Sainz, María; McGuigan, Kevin G; Ares-Mazás, Elvira
The solar disinfection (SODIS) technique is a highly effective process that makes use of solar energy to inactivate pathogenic microorganisms in drinking water in developing countries. The pathogenic protozoan parasite Cryptosporidium parvum is often found in surface waters and is associated with waterborne outbreaks of cryptosporidiosis. In the present study, a complete multi-factorial mathematical model was used to investigate the combined effects of the intensity of solar radiation (200, 600 and 900W/m(2) in the 320nm to 10microm range), water turbidity (5, 100 and 300 NTU) and exposure time (4, 8 and 12h) on the viability and infectivity of C. parvum oocysts during simulated SODIS procedures at a constant temperature of 30 degrees C. All three factors had significant effects (p or =600W/m(2) and times of exposure between 8 and 12h were required to reduce the oocyst infectivity in water samples with different degrees of turbidity.
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...
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...
Graczyk, T K; Cranfield, M R; Bostwick, E F
Therapy based on the protective passive immunity of hyperimmune bovine colostrum (HBC) was applied to 12 moribund Leopard geckos (Eublepharis macularius) infected with Cryptosporidium sp. The geckos were lethargic and moderately to severely emaciated, weighing on average 36% of the baseline body weight value. Seven gastric HBC treatments at 1-week intervals each decreased the relative output of Cryptosporidium sp. oocysts and the prevalence of oocyst-positive fecal specimens. Histologically, after 8 weeks of therapy, seven out of 12 geckos had only single developmental stages of Cryptosporidium sp. in the intestinal epithelium, and three, one and one geckos had low, moderate and high numbers, respectively, of the pathogen developmental stages. The HBC therapy was efficacious in decreasing the parasite load in moribund geckos. Morphometric and immunologic analysis of Cryptosporidium sp. oocyst isolates originating from Leopard geckos (E. macularius) demonstrated differences between gecko-derived oocyst isolates and isolates of C. serpentis recovered from snakes.
Jezkova, Jana; Horcickova, Michaela; Hlaskova, Lenka; Sak, Bohumil; Kvetonova, Dana; Novak, Jan; Hofmannova, Lada; McEvoy, John; Kvac, Martin
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
Yang, Rongchang; Murphy, Cain; Song, Yong; Ng-Hublin, Josephine; Estcourt, Annika; Hijjawi, Nawal; Chalmers, Rachel; Hadfield, Stephen; Bath, Andrew; Gordon, Cameron; Ryan, Una
Cryptosporidium is an enteric protozoan parasite that is resistant to inactivation by commonly used drinking water disinfectants. Between 2004 and 2010, it was responsible for 60% of all waterborne protozoan parasitic outbreaks reported worldwide. Most sporadic infections in humans and almost all outbreaks are caused by Cryptosporidium parvum and Cryptosporidium hominis. We report the development and validation of a quantitative qPCR assay using minor groove binder (MGB)-probes targeting a unique Cryptosporidium specific protein-coding gene, that directly detects, quantitates and identifies C. hominis and C. parvum in environmental and faecal samples. An internal amplification control (IAC) was also developed and included in this assay. The qPCR assay was compared with an 18S nested PCR assay for sensitivity and specificity. The analytical sensitivity for the qPCR assay was 1 oocyst and 1-10 oocysts for the 18S assay. Evaluation of analytical specificity of the qPCR assay revealed no cross-reactions with other genera and detected all C. parvum and C. hominis isolates correctly. The diagnostic sensitivity and specificity of the qPCR was 100% compared to 96.9% and 98.4%, respectively for the 18S assay. The qPCR assay was also highly reproducible with RSD (relative standard deviation) values of 1.4-9.4%, when the assay was performed by four different technicians. When tested on water samples, the qPCR assay was more sensitive than the 18S assay, detecting positives in 37 of 138 water samples compared to 35 for the 18S locus. This qPCR assay should be a valuable tool for the detection and differentiation of C. hominis and C. parvum in both clinical and environmental samples. Copyright © 2013 Elsevier Inc. All rights reserved.
Kik, Marja J L; van Asten, Alphons J A M; Lenstra, Johannes A; Kirpensteijn, Jolle
Cryptosporidium infection was associated with colitis and cystitis in 2 green iguanas (Iguana iguana). The disease was characterized by a chronic clinical course of cloacal prolapses and cystitis. Histological examination of the gut and urinary bladder showed numerous Cryptosporidium developmental stages on the surface of the epithelium with mixed inflammatory response in the lamina propria. Cryptosporidium oocysts were visualised in a cytological preparation of the faeces. Based on the small subunit ribosomal RNA gene the cryptosporidia were characterized as belonging to the intestinal cryptosporidial lineage, but not to Cryptosporidium saurophilum or Cryptosporidium serpentis species. Copyright © 2010 Elsevier B.V. All rights reserved.
Petersen, T. B.; Petersen, H. H.; Abaidoo, R. C.
Protozoan parasites belonging to the genus Cryptosporidium are transmitted e.g. by food and water and may cause severe diarrhoea, dehydration, weight loss and malnutrition. Ingestion of 10 oocysts can lead to infection and pathogenic symptoms. Thus, to characterize Cryptosporidium spp. contaminat...
Paziewska-Harris, A.; Schoone, G.; Schallig, H. D. F. H.
The long-term storage of Cryptosporidium life-cycle stages is a prerequisite for in vitro culture of the parasite. Cryptosporidium parvum oocysts, sporozoites, and intracellular forms inside infected host cells were stored for 6-12 mo in liquid nitrogen utilizing different cryoprotectants (dimethyl
Le, Xiao-hua; Wang, Hui; Gou, Ji-zhou; Chen, Xin-chun; Yang, Gui-lin; Yang, Qian-ting; Li, Xiao-he; Zhou, Bo-ping; Li, Hui-qin; Cai, Wei-ping
To investigate the infection of Cryptosporidium and its epidemiological characteristics in AIDS patients of Southern China. Stool samples colleted from AIDS confirmed patients. The samples were detected for oocyst of Cryptosporidium by acid fast bacteria stain and indirect fluorescent antibody stain respectively, CD4 count was detected by Flow Cytometry. 212 samples of fresh stool obtained from the AIDS patients who live in Guangdong and Yunnan province. The total infection rate of Cryptosporidium in AIDS patients was 4.25% (9/212), the infectious rate of oocyst in the group of 50- 59-years-old was significantly higher than those in 30-39 (P stool shape. AIDS patients infected by Cryptosporidium are not rare in southern China, and the infectious rate was lower than western country. Patients received ART could decrease the infectious rate of Cryptosporidium, Cryptosporidium always happen in patient whose CD4 count was very low (< 100 cells/microl).
To secure the safety and quality of food and water presence of pathogens including parasites must be controlled. This relies among other things on implementation of sensitive and specific tests which are able to rule out the occurrence of parasites. Worldwide environmentally resistant oocysts and cysts ((oo-)cysts) of the protozoan genera Cryptosporidium and Giardia are major causes of waterborne outbreaks of diarrhea. Methods for routine recovery and detection of waterborne Giardia and/or Cr...
Full Text Available Of 1 346 faecal samples from the Chikwawa and Thyolo districts of Malawi, analysed for the presence of Cryptosporidium oocysts between October 2001 and May 2003, 61.3 % were from cattle (29.8 % of these were from calves < 6 months old. Cryptosporidium oocysts were detected during all three seasons studied in Chikwawa and Thyolo. In Chikwawa, 13.6 % of adult cattle and 11.7 % of calves were infected, compared to 28.9 % of adult cattle and 36.7 % of calves in Thyolo. Dependent on season, between 7.8 % and 37.7 % (Chikwawa and 16.7 % and 39.3 % (Thyolo of cattle samples contained oocysts. In Chikwawa, the highest percentage of infections occurred in the cool season, whereas in Thyolo, the highest percentage of infections occurred in the dry season. Faecal samples from goats [n = 225], pigs [n = 92], sheep [n = 6], rabbits, guinea pigs, chickens, ducks, turkeys, doves and guinea fowls were also analysed. Up to 5.6 % of goat samples contained oocysts in Chikwawa, compared to between 16.7 % and 39.3 % in Thyolo. Again, in Chikwawa, the highest percentage of infections occurred in the cool season and the lowest in the rainy season, whereas, in Thyolo, the highest percentage of infections occurred in the dry season and the lowest in the cool season. In pigs, more infections were detected in the dry season in Chikwawa, but infections in the cool season were similar (17.7 %, whereas in Thyolo, infections occurred in all three seasons (17.9 % in the rainy season, 25 % in the cool season and 60 % in the dry season. Often diarrhoeic, oocyst positive cattle faecal samples collected from Chikwawa and subjected to PCR-RFLP, four oocyst positive samples (two from heifers, one from a cow and one unknown were amplified at an 18S rRNA and Cryptosporidium oocyst wall protein (COWP loci. RFLP of the 18S rRNA locus indicated that Cryptosporidium parvum, Cryptosporidium hominis, Cryptosporidium bovis and / or Cryptosporidium ryanae DNA, or a mixture of them was present
Widerström, Micael; Schönning, Caroline; Lilja, Mikael; Lebbad, Marianne; Ljung, Thomas; Allestam, Görel; Ferm, Martin; Björkholm, Britta; Hansen, Anette; Hiltula, Jari; Långmark, Jonas; Löfdahl, Margareta; Omberg, Maria; Reuterwall, Christina; Samuelsson, Eva; Widgren, Katarina; Wallensten, Anders; Lindh, Johan
In November 2010, ≈27,000 (≈45%) inhabitants of Östersund, Sweden, were affected by a waterborne outbreak of cryptosporidiosis. The outbreak was characterized by a rapid onset and high attack rate, especially among young and middle-aged persons. Young age, number of infected family members, amount of water consumed daily, and gluten intolerance were identified as risk factors for acquiring cryptosporidiosis. Also, chronic intestinal disease and young age were significantly associated with prolonged diarrhea. Identification of Cryptosporidium hominis subtype IbA10G2 in human and environmental samples and consistently low numbers of oocysts in drinking water confirmed insufficient reduction of parasites by the municipal water treatment plant. The current outbreak shows that use of inadequate microbial barriers at water treatment plants can have serious consequences for public health. This risk can be minimized by optimizing control of raw water quality and employing multiple barriers that remove or inactivate all groups of pathogens.
Hofstra, N; Bouwman, A F; Beusen, A H W; Medema, G J
The protozoan parasite Cryptosporidium is a major cause of diarrhoea worldwide. This paper presents the first model-based inventory with 0.5 by 0.5 degree resolution of global Cryptosporidium emissions for the year 2000 from humans and animals to surface water. The model is based on nutrient distribution modelling, because the sources and transport of oocysts and nutrients to the surface water are comparable. Total emissions consist of point source emissions from wastewater and nonpoint source emissions by runoff of oocysts in manure from agricultural lands. Results indicate a global emission of 3 × 10(17) oocysts per year, with comparable contributions from point and nonpoint sources. Hot-spot areas for point sources are big cities in China, India and Latin America, while the area with the largest nonpoint source emissions is in China. Uncertainties in the model are large. Main areas for further study are (i) excretion rates of oocysts by humans and animals, (ii) emissions of humans not connected to sewage systems, and (iii) retention of oocysts to determine surface water pathogen concentrations rather than emissions. Our results are useful to health organisations to identify priority areas for further study and intervention. Copyright © 2012 Elsevier B.V. All rights reserved.
Ping Lu; Tao Yuan; Baojun Zhang
Transport of Cryptosporidium in runoff will contaminate the surrounding water body. Cryptosporidium-sized microspheres and inactive Cryptosporidium parvum oocysts removal using an active carbon (AC) filter with/without coal gangue from the runoff was evaluated. Chemical composition of coal gangue was determined, and its performance as a water treatment material was tested. Results showed SiO2 was the dominant chemical component of coal gangue. Removal was only 20% for microspheres and 24% for...
Li, Juan; Lin, Xuhui; Zhang, Longxian; Qi, Nanshan; Liao, Shenquan; Lv, Minna; Wu, Caiyan; Sun, Mingfei
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.
Kvác, Martin; Hofmannová, Lada; Bertolino, Sandro; Wauters, Luc; Tosi, Guido; Modrý, David
We investigated the genotypes of Cryptosporidium infecting red squirrels (Sciurus vulgaris L.) in two areas of the Western Alps in Italy. Examination of 141 faecal samples from 70 red squirrels revealed oocysts of Cryptosporidium in 17 animals (24.3%). Based on 18S rRNA gene sequencing, two genotypes of Cryptosporidium species were found: 15 squirrels were positive for the Cryptosporidium ferret genotype and 2 for the Cryptosporidium chipmunk genotype I. The occurrence and intensity of Cryptosporidium infection did not differ between the study areas or sex. More than 85% of the positive animals were adults; however no difference was found between Cryptosporidium infection in the juvenile and adult age groups. Oocysts of the Cryptosporidium ferret genotype measured 5.5 +/- 0.3 x 5.2 +/- 0.2 microm (shape index 1.06) and the Cryptosporidium chipmunk genotype I 5.8 +/- 0.3 x 5.4 +/- 0.3 microm (shape index 1.07). Neonatal and adult CD1 and BABL/c mice inoculated with 1 x 10(3) fresh oocysts of both genotypes did not produce detectable infection.
Castro-Hermida, José Antonio; González-Warleta, Marta; Mezo, Mercedes
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. Copyright © 2014 Elsevier GmbH. All rights reserved.
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.
Dellarupe, A; Unzaga, J M; Moré, G; Kienast, M; Larsen, A; Stiebel, C; Rambeaud, M; Venturini, M C
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.
Yimming, Benjarat; Pattanatanang, Khampee; Sanyathitiseree, Pornchai; Inpankaew, Tawin; Kamyingkird, Ketsarin; Pinyopanuwat, Nongnuch; Chimnoi, Wissanuwat; Phasuk, Jumnongjit
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.
... to be effective. If you have a weakened immune system, ask your provider if you need to boil your water. Alternative Names Cryptosporidiosis Images Cryptosporidium, organism Digestive system organs References Huston CD. Intestinal protozoa. In: ...
Pagoso, Edison Jay A; Rivera, Windell L
Manila Bay is one of the major propagation sites of edible bivalves in the Philippines. Studies have shown that bivalves might be contaminated with human pathogens like the protozoan parasite Cryptosporidium, one of the major causes of gastroenteritis in the world. In this study, Cryptosporidium from four species of edible bivalves were isolated using a combination of sucrose flotation and immunomagnetic separation. Using direct fluorescent antibody test, Cryptosporidium oocysts were found in 67 out of 144 samples collected. DNA sequence analysis of the 18S rRNA gene of the isolates detected C. parvum and C. hominis (major causes of human cryptosporidiosis) and C. meleagridis (causes infection in avian species). Analysis of the 60kDa glycoprotein gene further confirmed the genotypes of the Cryptosporidium isolates. This study is the first to provide baseline information on Cryptosporidium contamination of Manila Bay where bivalves are commonly cultured. Copyright © 2017 Elsevier Ltd. All rights reserved.
Hijnen, W.A.M.; Visser, Ate; Schijven, J.F.; Bonné, P.; Medema, Gerriet Jan
The decimal elimination capacity (DEC) of slow sand filters (SSF) for viruses, bacteria and oocysts of Cryptosporidium has been assessed from full-scale data and pilot plant and laboratory experiments. DEC for viruses calculated from experimental data with MS2-bacteriophages in the pilot plant
Wang, Rongjun; Wang, Jinchan; Sun, Mingfei; Dang, Hailiang; Feng, Yaoyu; Ning, Changshen; Jian, Fuchun; Zhang, Longxian; Xiao, Lihua
A total of 124 fecal specimens were collected from four deer farms in Zhengzhou City, China and examined for Cryptosporidium by Sheather's sugar flotation technique. Cryptosporidim oocysts were detected in two 1-year-old sika deer, and one of the two specimens was genotyped by sequence and phylogenetic analyses of the small subunit ribosomal RNA (rRNA) (18S rRNA), 70-kDa heat shock protein (HSP70), actin, and Cryptosporidium oocyst wall protein (COWP) genes. Results obtained suggested that the Cryptosporidium studied belonged to Cryptosporidium cervine genotype, although slight sequence differences were noticed at the three loci. The similarities between this isolate and other Cryptosporidium cervine genotype isolates were 99.1-99.8%, 9.8%, 99.7%, and 100% at the 18S rRNA, HSP70, actin, and COWP loci, respectively. This study is the first report of Cryptosporidium infection in sika deer in China.
Chalmers, Rachel M; Robinson, Guy; Elwin, Kristin; Hadfield, Stephen J; Thomas, Euron; Watkins, John; Casemore, David; Kay, David
As part of investigations into the cause of a waterborne outbreak of Cryptosporidium hominis infection linked to a mains water supply, surface waters and wastewater treatment plants were tested for Cryptosporidium spp. Oocyst counts in base flow surface water samples ranged from nil to 29 per 10 l. Oocyst counts in effluent from a community wastewater treatment plant were up to 63 fold higher and breakout from one septic tank five logs higher. There were no peak (storm) flow events during the investigation. C. hominis, four named genotypes (cervine, muskrat II, rat, W19) and six new small subunit ribosomal RNA gene sequences were identified. Four of the new sequences were closely related to Cryptosporidium muskrat genotype I, one was closely related to the fox genotype and one to Cryptosporidium canis. C. hominis was found extensively in the catchment, but only at sites contaminated by wastewater, and in the treated water supply to the affected area. All were gp60 subtype IbA10G2, the outbreak subtype. Multiple routes of contamination of the reservoir were identified, resulting in persistent detection of low numbers of oocysts in the final water. This work demonstrates the utility of genotyping Cryptosporidium isolates in environmental samples during outbreak investigations.
Niine, Tarmo; Dorbek-Kolin, Elisabeth; Lassen, Brian
Cryptosporidium spp. infections in neonatal dairy calves can cause diarrhoea and, in rare cases, death. The infection is usually self-limiting, but halofuginone lactate (HL) can be used prophylactically. Calves (n = 144) in the study were born during a 2-month period on one farm. A total of 901...... of life. HP concentration and HL treatment were negatively associated with weight gain at 3 months of age. Cryptosporidium positive faecal samples were significantly (P ... the shedding of Cryptosporidium oocysts and improved survival, but was negatively associated with weight gain. Incorrect treatment had a low impact on mortality and resembled no treatment regarding the proportion of calves shedding oocysts. Acute phase response (APR) in the second week of life seemed...
Araújo, Ronalda S; Dropa, Milena; Fernandes, Licia N; Carvalho, Terezinha T; Sato, Maria Inês Z; Soares, Rodrigo M; Matté, Glavur R; Matté, Maria Helena
The protozoan parasite Cryptosporidium has emerged as one of the most important water contaminants, causing waterborne outbreaks of diarrheal diseases worldwide. The small size of oocysts under the microscope and the possibility of changes in characteristics of oocysts, mainly in environmental samples, make the taxonomy of the genus difficult if morphologic characteristics are considered. This limitation encouraged the application of molecular methods to identify this microorganism. The aim of this study was to detect and identify by nested-polymerase chain reaction oocysts of Cryptosporidium present in water samples in the state of São Paulo, Brazil. Water samples were concentrated through a membrane filter, DNA was extracted by using a standard technique, and both amplification reactions used forward and reverse oligonucleotides that were complementary to Cryptosporidium 18S ribosomal RNA gene sequences. Thirty water samples from different sites of collection in the state of São Paulo were evaluated. Cryptosporidium oocysts were detected in 30% of the samples. By genoptyping, C. hominis and Cryptosporidium sp. were identified in recreational water and C. meleagridis was identified in surface water samples. This is the first report of C. hominis in environmental samples in Brazil. Although identification of Cryptosporidium is still a difficult task, molecular methods are essential for specific identification and are a helpful tool to aid to understand the epidemiology of this parasite in Brazil.
Heaselgrave, Wayne; Kilvington, Simon
The antimicrobial activity of simulated solar disinfection (SODIS) in the presence and absence of riboflavin against various protozoa and helminth organisms was investigated in this study. Assays were conducted in transparent 12 well microtitre plates containing a suspension of test organisms in the presence or absence of 250 μM riboflavin. Plates were exposed to simulated sunlight at an optical irradiance of 550 Wm(-2) (watts per square metre) delivered from a SUNTEST™ CPS+ solar simulator. Aliquots of the test suspensions were taken at set time points and the viability of the test organisms was determined by either culture, microscopy or flow cytometry where applicable. With Acanthamoeba, Naegleria, Entamoeba and Giardia exposure to SODIS at an optical irradiance of 550 Wm(-2) for up to 6h resulted in significant inactivation of these organisms. The addition of riboflavin to this system significantly increased the level of inactivation observed with cysts of A. castellanii. With Cryptosporidium oocysts and Ascaris ova exposure to SODIS in the presence and absence of riboflavin for 6-8h resulted in a negligible reduction in viability of both organisms. In this present study we have been able to show that SODIS is effective against a variety of previously untested waterborne organisms and with A. castellanii cysts the addition of micro-molar concentrations of riboflavin can enhance cyst inactivation. However, care must be taken as Ascaris larvae continue to develop inside the ova after exposure to SODIS and Cryptosporidium remain impermeable to propidium iodide staining indicating they may still be infectious. Copyright © 2011 Elsevier B.V. All rights reserved.
Gu, Youfang; Wang, Xiaolong; Zhou, Cefan; Li, Peiying; Xu, Qianming; Zhao, Changcheng; Liu, Wei; Xu, Wenlong
To assess Cryptosporidium infections among wild animals in a zoo located in Anhui province, we conducted an investigation on the fecal samples collected from 44 primates, 41 herbivores, 44 carnivores and omnivores, and 103 birds in the zoo with the use of Sheather's sugar flotation technique and modified acid-fast staining. Cryptosporidium oocysts were detected in the fecal samples from six primates, two herbivores, four carnivores and omnivores, and seven birds by using Sheather's sugar flotation technique; the prevalence of Cryptosporidium infection in primates, herbivores, carnivores and omnivores and birds was 13.64, 4.88, 9.09, and 6.80%, respectively. Modified acid-fast staining detected the presence of Cryptosporidium oocysts in the fecal samples of one primate, three herbivores, 0 carnivores and omnivores, and one bird, and the prevalence of Cryptosporidium infection in primates, herbivores, carnivores and omnivores and birds was 2.27, 7.32, 0.00, and 0.97%, respectively. Polymerase chain reaction-restriction fragment-length polymorphism (PCR-RFLP) and phylogenetic analysis with the use of the neighbor-joining (NJ) method based on the aligned partial small-subunit (SSU) rRNA gene sequences showed that the protozoan pathogen isolated from primates was Cryptosporidium hominis and the pathogen isolated from camels ( Camelus dromedarius ) was Cryptosporidium andersoni. Subtyping the Cryptosporidium hominis by 60-kDa glycoprotein (GP60) gene phylogenetic analysis showed the Cryptosporidium hominis belongs to the subtype IdA and IbA.
Hijnen, W.A.M.; Veer, A.J. van der; Beveren, J. van; Medema, Gerriet Jan
The inactivation of C. parvum and spores of C. perfringens by ozone treatment in natural water was compared in a lab-scale continuous-flow system. In addition the inactivation of the natural occurring spores of sulphite-reducing clostridia (SSRC) in this water was monitored in one of the lab-scale
Full Text Available Purpose: The aim of the present study was to determine the prevalence of Crytposporidium in local population and to understand its epidemiology by molecular methods. Methods: Faecal samples from 681 children and 804 adults, admitted to tertiary care hospitals in twin cities of Hyderabad and Secunderabad with complaints of diarrhoea; and six calves with diarrhoea, were screened for Cryptosporidium oocysts by microscopy and enzyme linked immunosorbent assay (ELISA. Polymerase chain reaction restriction fragment length polymorphism (PCR RFLP based identification of Cryptosporidium species in positive specimens was done to elucidate epidemiology of Cryptosporidium. Results : Cryptosporidium was found in 52 (7.6% children and 7(0.9% adults and 1(16.6% calf with diarrhoea. The prevalence of Cryptosporidium in children below five years of age was 8.2% and 14.3% in children in the age group of six months to one year. Of the 42 samples genotyped 29 (69% were C. hominis and 8 (19% were C. parvum and 5 (11.9% were mixed infection with the two species. Conclusions : Children in the age group of six months to one year were found to be the most vulnerable. The occurrence of C. parvum , in nearly one third of cases in the present series indicates that the zoonotic transmission is of considerable significance in the epidemiology of Cryptosporidiosis in the study area.
of immunofluorescence stained (oo-)cysts. These methods have low recovery rates, are time consuming, costly, and require well equipped laboratory facilities. Likewise, microscopy is the universal diagnostic method for detection of helminth eggs and protozoa in food and feed despite low sensitivity, difficulties......To secure the safety and quality of food and water presence of pathogens including parasites must be controlled. This relies among other things on implementation of sensitive and specific tests which are able to rule out the occurrence of parasites. Worldwide environmentally resistant oocysts...... system and ultrasound to obtain high recovery rates of apparently undamaged protozoa: 84.9% (Standard deviation (±) 4.8) for Giardia cysts and 70% (± 6.5) for Cryptosporidium oocysts. Ultrasound in the current system is tuned into a useful tool for enhanced elution of filtered (oo-)cysts. The combined...
Bushen, Oluma Y; Kohli, Anita; Pinkerton, Relana C; Dupnik, Kate; Newman, Robert D; Sears, Cynthia L; Fayer, Ronald; Lima, Aldo A M; Guerrant, Richard L
Cryptosporidium is an important cause of infectious diarrhoea worldwide, but little is known about the course of illness when infected with different species. Over a period of 5 years, Cryptosporidium was identified in the stools of 58 of 157 children prospectively followed from birth in an urban slum (favela) in northeast Brazil. Forty isolates were available for quantification and 42 for speciation (24 Cryptosporidium hominis and 18 C. parvum). Children with C. hominis shed significantly more oocysts/ml of stool (3.5 x 10(6) vs. 1.7 x 10(6)perml; P=0.001), and oocyst counts were higher among symptomatic children (P=0.002). Heavier C. parvum shedding was significantly associated with symptoms (P=0.004), and symptomatic C. parvum-infected children were significantly more likely than asymptomatic children to be lactoferrin-positive (P=0.004). Height-for-age (HAZ) Z-scores showed significant declines within 3 months of infection for children infected with either C. hominis (P=0.028) or C. parvum (P=0.001). However, in the 3-6 month period following infection, only C. hominis-infected children continued to demonstrate declining HAZ score and asymptomatic children showed even greater decline (P=0.01). Cryptosporidium hominis is more common than C. parvum in favela children and is associated with heavier infections and greater growth shortfalls, even in the absence of symptoms.
Sotiriadou, Isaia; Pantchev, Nikola; Gassmann, Doreen; Karanis, Panagiotis
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. PMID:23477297
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.
Rinaldi, L; Capasso, M; Mihalca, A D; Cirillo, R; Cringoli, G; Cacciò, S
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).
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.
Zhang, Xiao-Ping; He, Yan-Yan; Zhu, Qian; Ma, Xiao-Jiang; Cai, Li
To understand the contamination status of Cryptosporidium sp. and Giardia lamblia in drinking water, source water and environmental water in Shanghai. All water samples collected from drinking water, source water and environmental water were detected by a procedure of micromembrane filtration, immune magnetic separation (IMS), and immunofluorescent assay (IFA). Cryptosporidium oocysts and Giardia cysts were not found in 156 samples of the drinking water including finished water, tap water, or pipe water for directly drinking in communities. Among 70 samples either source water of water plants (15 samples), environmental water from Huangpu River(25), canal water around animal sheds(15), exit water from waste-water treatment plants(9), or waste water due to daily life(6), Cryptosporidium oocysts were detected in 1(6.7%), 2(8.0%), 7(46.7%), 1(11.1%), and 1(16.7%) samples, respectively; and Giardia cysts were detected in 1(6.7%), 3(12.0%), 6 (40.0%), 2(22.2%), and 2(33.3%), respectively. The positive rate of Cryptosporidium oocysts and Giardia cysts was 17.1% (12/70) and 20.0% (14/70), respectively. No Cryptosporidium oocysts and Giardia cysts have been detected in drinking water, but found in source water and environmental water samples in Shanghai.
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
Lv, Chaochao; Zhang, Longxian; Wang, Rongjun; Jian, Fuchun; Zhang, Sumei; Ning, Changshen; Wang, Helei; Feng, Chao; Wang, Xinwei; Ren, Xupeng; Qi, Meng; Xiao, Lihua
To understand the prevalence of Cryptosporidium infection in rodents in China and to assess the potential role of rodents as a source for human cryptosporidiosis, 723 specimens from 18 rodent species were collected from four provinces of China and examined between August 2007 and December 2008 by microscopy after using Sheather's sugar flotation and modified acid-fast staining. Cryptosporidium oocysts were detected in 83 specimens, with an overall prevalence of 11.5%. Phodopus sungorus, Phodo...
Rimhanen-Finne, R.; Enemark, Heidi L.; Kolehmainen, J.
The performance of immunofluorescence microscopy (IF) and enzyme-linked immunosorbent assay (ELISA) in canine feces was evaluated. IF and Cryptosporidium ELISA detected 10(5) oocysts/g, while the detection limit for Giardia ELISA was 10(4) cysts/g. The Cryptosporidium ELISA showed 94% specificity...... zoonotic character of Cryptosporidium and Giardia infections in 150 asymptomatic Finnish dogs from the Helsinki area were studied. The overall proportion of dogs positive for Cryptosporidium was 5% (7/150) and that for Giardia 5% (8/150). In dogs...
Yang, Wenli; Chen, Plato; Villegas, Eric N.; Landy, Ronald B.; Kanetsky, Charles; Cama, Vitaliano; Dearen, Theresa; Schultz, Cherie L.; Orndorff, Kenneth G.; Prelewicz, Gregory J.; Brown, Miranda H.; Young, Kim Roy; Xiao, Lihua
To better characterize Cryptosporidium in the Potomac River watershed, a PCR-based genotyping tool was used to analyze 64 base flow and 28 storm flow samples from five sites in the watershed. These sites included two water treatment plant intakes, as well as three upstream sites, each associated with a different type of land use. The uses, including urban wastewater, agricultural (cattle) wastewater, and wildlife, posed different risks in terms of the potential contribution of Cryptosporidium oocysts to the source water. Cryptosporidium was detected in 27 base flow water samples and 23 storm flow water samples. The most frequently detected species was C. andersoni (detected in 41 samples), while 14 other species or genotypes, almost all wildlife associated, were occasionally detected. The two common human-pathogenic species, C. hominis and C. parvum, were not detected. Although C. andersoni was common at all four sites influenced by agriculture, it was largely absent at the urban wastewater site. There were very few positive samples as determined by Environmental Protection Agency method 1623 at any site; only 8 of 90 samples analyzed (9%) were positive for Cryptosporidium as determined by microscopy. The genotyping results suggest that many of the Cryptosporidium oocysts in the water treatment plant source waters were from old calves and adult cattle and might not pose a significant risk to human health. PMID:18776033
Ciçek, Mutalip; Körkoca, Hanifi; Gül, Abdurrahman
This study was carried out in order to investigate the prevalence of Cryptosporidium sp. in slaughtered animals and workers of the Van municipality slaughterhouse in Van. Animals slaughtered at different times and workers who had been working in different departments of the slaughter house were included in the study for three months. A total of 309 fecal specimens from animals including 167 sheep, 56 goats and 86 cattle and 87 fecal specimens from workers were examined for Cryptosporidium sp. oocysts. In slaughtered animals, the modified acid-fast staining method was used to determine the oocysts of Cryptosporidium sp. The fecal samples of slaughter workers were examined by using RIDA (R) Quick Cryptosporidium Strip Test (R-Biopharm, Germany) and the modified acid-fast staining method. Fecal samples found to be positive by stripe test were also confirmed with the ELISA method (R-Biopharm, Germany). Oocysts of Cryptosporidium sp. were found in fecal specimens of 22 sheep (13.17%), 6 goats (10.71%) and 7 cattle (8.13%). Intestinal parasites were observed in 34 fecal specimens of workers (39.08%). Cryptosporidium sp., Hymenolepis nana, Chilomastix mesnili, Endolimax nana, Iodamoeba bütschlii were found in the specimen of one worker (1.14%), Entamoeba coli in 4 workers (4.59%), Blastocystis hominis (9.19%) in 8 workers, and Giardia intestinalis (19.54%) in 17 workers.
N Taghipour Lailabadi
Full Text Available Background: The protozoan parasites Cryptosporidium spp. and Giardia are known to occur widely in both raw and drinking waters. They are two of the causative agents of waterborne out-breaks of gastroenteritis throughout the world. In the present study, a PCR assay and FA were developed for detection of Cryptosporidium oocysts and Giardia cyst in environmental samples. Methods: We have detected Cryptosporidium spp. oocysts and Giardia cysts in seeded and un-seeded environmental water samples by PCR method. Water samples were spiked with oocysts (50, 100,300,500 and filtrated with a 1.2-µm pore size cellulose nitrate and follow by DNA extrac¬tion and purification by QIAamp DNA mini kit. Nested-PCR assay amplified an 850 bp fragment of 18s rRNA gene specific for Cryptosporidium and 435 bp fragment of glutamate dehydrogenase (GDH target gene for Giardia. Also many river water from north of Iran, be checked by these methods. Results: Cryptosporidium and Giardia DNAs were detected in seeded water sample and Giardia was detected in all 5 water samples from river in north of Iran by nested- PCR and FA. Also in one river water sample, Cryptosporidium was detected.Conclusion: This protocol is effective for detection of these waterborne parasites in treated and untreated water samples. This study can also serve as a platform for further investigations and research water source in Iran.
McAllister, Chris T.; Duszynski, Donald W.; Fisher, Robert N.
Between 1991 and 1993, 295 lizards, comprising 21 species in 2 families (Gekkonidae, Scincidae) from the Cook Islands, Fiji, Palau, Takapoto, and Vanuatu in the South Pacific, were examined for Cryptosporidium oocysts. Only 6 lizards (2%) were found to be passing Cryptosporidium oocysts in their feces, including 2 of 30 (7%) Oceania geckos, Gehyra oceanica, from Rarotonga, Cook Islands, and 4 of 26 (15%) Pacific blue-tailed skinks, Emoia caeruleocauda, from Efate Island, Vanuatu. This represents the largest survey for Cryptosporidium in Pacific island lizards, and we document 2 new host and 2 new locality records for this parasite genus.
Full Text Available The protozoan intestinal parasite Cryptosporidium commonly infects cattle throughout the world and Iran. The present study was undertaken to determine the abundance and associated risk factors of Cryptosporidium infection in cattle herds of northwestern Iran. A total number of 246 fecal samples from 138 (56.1% diarrheic (D and 108 (43.9% non-diarrheic (ND cattle were randomly collected and examined by fecal smears stained with Ziehl-Neelsen. For molecular specification, DNA was extracted from collected Cryptosporidium oocysts and a fragment of 1325 bp in size from 18S rRNA gene was amplified. The overall prevalence of Cryptosporidium infection was 22.3% (55/246. The prevalence of Cryptosporidium infection in examined calves less than 6 month-old was significantly higher than adult cattle. C. parvum and C. andersoni were identified in 20.3% (50/246 and 2.03% (5/246 of examined cattle, respectively. The highest prevalence of C. parvum infection was found in D calves < 6 month-old (13.4%, 33/246, while C. andersoni was only detected in ND cattle (8.9%, 22/246. There was significant difference in the prevalence between male than female cattle. There was no significant difference between prevalence and seasons of investigation. It was concluded that C. parvum was the prevalent species in younger animals compared to older ones as a potentially zoonotic agent in the region.
Hadfield, Stephen J; Pachebat, Justin A; Swain, Martin T; Robinson, Guy; Cameron, Simon Js; Alexander, Jenna; Hegarty, Matthew J; Elwin, Kristin; Chalmers, Rachel M
Whole genome sequencing (WGS) of Cryptosporidium spp. has previously relied on propagation of the parasite in animals to generate enough oocysts from which to extract DNA of sufficient quantity and purity for analysis. We have developed and validated a method for preparation of genomic Cryptosporidium DNA suitable for WGS directly from human stool samples and used it to generate 10 high-quality whole Cryptosporidium genome assemblies. Our method uses a combination of salt flotation, immunomagnetic separation (IMS), and surface sterilisation of oocysts prior to DNA extraction, with subsequent use of the transposome-based Nextera XT kit to generate libraries for sequencing on Illumina platforms. IMS was found to be superior to caesium chloride density centrifugation for purification of oocysts from small volume stool samples and for reducing levels of contaminant DNA. The IMS-based method was used initially to sequence whole genomes of Cryptosporidium hominis gp60 subtype IbA10G2 and Cryptosporidium parvum gp60 subtype IIaA19G1R2 from small amounts of stool left over from diagnostic testing of clinical cases of cryptosporidiosis. The C. parvum isolate was sequenced to a mean depth of 51.8X with reads covering 100 % of the bases of the C. parvum Iowa II reference genome (Bioproject PRJNA 15586), while the C. hominis isolate was sequenced to a mean depth of 34.7X with reads covering 98 % of the bases of the C. hominis TU502 v1 reference genome (Bioproject PRJNA 15585). The method was then applied to a further 17 stools, successfully generating another eight new whole genome sequences, of which two were C. hominis (gp60 subtypes IbA10G2 and IaA14R3) and six C. parvum (gp60 subtypes IIaA15G2R1 from three samples, and one each of IIaA17G1R1, IIaA18G2R1, and IIdA22G1), demonstrating the utility of this method to sequence Cryptosporidium genomes directly from clinical samples. This development is especially important as it reduces the requirement to propagate
Lv, Chaochao; Zhang, Longxian; Wang, Rongjun; Jian, Fuchun; Zhang, Sumei; Ning, Changshen; Wang, Helei; Feng, Chao; Wang, Xinwei; Ren, Xupeng; Qi, Meng; Xiao, Lihua
To understand the prevalence of Cryptosporidium infection in rodents in China and to assess the potential role of rodents as a source for human cryptosporidiosis, 723 specimens from 18 rodent species were collected from four provinces of China and examined between August 2007 and December 2008 by microscopy after using Sheather's sugar flotation and modified acid-fast staining. Cryptosporidium oocysts were detected in 83 specimens, with an overall prevalence of 11.5%. Phodopus sungorus, Phodopus campbelli, and Rattus tanezumi were new reported hosts of Cryptosporidium. The genotypes and subtypes of Cryptosporidium strains in microscopy-positive specimens were further identified by PCR and sequence analysis of the small subunit rRNA and the 60-kDa glycoprotein (gp60) genes. In addition to Cryptosporidium parvum, C. muris, C. andersoni, C. wrairi, ferret genotype, and mouse genotype I, four new Cryptosporidium genotypes were identified, including the hamster genotype, chipmunk genotype III, and rat genotypes II and III. Mixed Cryptosporidium species/genotypes were found in 10.8% of Cryptosporidium-positive specimens. Sequence analysis of the gp60 gene showed that C. parvum strains in pet Siberian chipmunks and hamsters were all of the subtype IIdA15G1, which was found previously in a human isolate in The Netherlands and lambs in Spain. The gp60 sequences of C. wrairi and the Cryptosporidium ferret genotype and mouse genotype I were also obtained. These findings suggest that pet rodents may be potential reservoirs of zoonotic Cryptosporidium species and subtypes.
Enemark, Heidi L.; Ahrens, Peter; Juel, Cynthia Dawn
The genetic polymorphism among 271 Danish Cryptosporidium isolates of human and animal origin was studied by partial amplification and sequencing of the Cryptosporidium oocyst wall protein (COWP) gene, the 18S rDNA, and a microsatellite locus.dagger Furthermore, the microsatellite locus was studied...... directly using fragment analysis. A comparative analysis of DNA sequences showed the presence of 3 different subgenotypes (C1, C2 and C3) in C. parvum isolates from Danish cattle, with prevalences of 16.7, 17.2 and 73.1% including 13 (7.0%) mixed infections. Subgenotype C1 was significantly more prevalent...... (P isolates of human origin the anthroponotic subgenotype H1 was identified, in addition to the zoonotic subgenotypes C1, C2, and C3. Of 44 human samples, 56.8% were anthroponotic, whereas 40.9% were zoonotic genotypes. One human isolate...
Smith, H V; Paton, C A; Mitambo, M M; Girdwood, R W
Cyclospora sp. oocysts sporulated maximally at 22 and 30 degrees C for 14 days retarded sporulation. Up to 12% of human- and baboon-derived oocysts previously stored at 4 degrees C for 1 to 2 months sporulated when stored for 6 to 7 days at 30 degrees C.
Full Text Available The prevalence of Cryptosporidium spp. infection in a cross-sectional study of dairy cattle, from two contrasting dairying regions in Tanzania, were determined by staining smears of faecal samples with the modified Ziehl-Neelsen technique. Of the 1 126 faecal samples screened, 19.7% were positive for Cryptosporidium spp. The prevalence was lower in Tanga Region than in Iringa Region. The prevalence of affected farms was 20% in Tanga and 21% in Iringa. In both regions, the probability of detecting Cryptosporidium oocysts in faeces varied with animal class, but these were not consistent in both regions. In Tanga Region, Cryptosporidium oocysts were significantly more likely to be found in the faeces of milking cows. In Iringa Region, the likelihood that cattle had Cryptosporidium-positive faeces declined with age, and milking cattle were significantly less likely to have Cryptosporidium positive faeces. In this region, 7% of cattle were housed within the family house at night, and this was marginally associated with a higher likelihood that animals had Cryptosporidium-positive faeces. Our study suggests that even though herd sizes are small, Cryptosporidium spp. are endemic on many Tanzanian smallholder dairy farms. These protozoa may impact on animal health and production, but also on human health, given the close associations between the cattle and their keepers. Further studies are required to assess these risks in more detail, and understand the epidemiology of Cryptosporidium spp. in this management system.
Bandyopadhyay, Kakali; Kellar, Kathryn L; Moura, Iaci; Casaqui Carollo, Maria Cristina; Graczyk, Thaddeus K; Slemenda, Susan; Johnston, Stephanie P; da Silva, Alexandre J
Cryptosporidium hominis and Cryptosporidium parvum are associated with massive disease outbreaks worldwide. Because these two species have different transmission cycles, identification of these parasites to the species level in clinical samples may provide laboratory data of crucial importance in epidemiologic investigations. To date, the most reliable way to differentiate C. hominis and C. parvum is based on DNA sequencing analysis of PCR amplicons. Although this approach is very effective for differentiation of Cryptosporidium species, it is labor-intensive and time-consuming compared with methods that do not require DNA sequencing analysis as an additional step and that have been successfully used for specific identification of a number of pathogens. In this study, we describe a novel Luminex-based assay that can differentiate C. hominis from C. parvum in a rapid and cost-effective manner. The assay was validated by testing a total of 143 DNA samples extracted from clinical specimens, environmental samples, or samples artificially spiked with Cryptosporidium oocysts. As few as 10 oocysts per 300 microl of stools could be detected with this assay. The assay format includes species-specific probes linked to carboxylated Luminex microspheres that hybridize to a Cryptosporidium microsatellite-2 region (ML-2) where C. hominis and C. parvum differ by one nucleotide substitution. The assay proved to be 100% specific when samples that had been characterized by direct fluorescent antibody test (DFA) and DNA sequencing analysis were tested. In addition, the assay was more sensitive than DFA and provided species identification, which is an advantage for epidemiologic studies.
Full Text Available Water is a vehicle for disseminating human and veterinary toxoplasmosis due to oocyst contamination. Several outbreaks of toxoplasmosis throughout the world have been related to contaminated drinking water. We have developed a method for the detection of Toxoplasma gondii oocysts in water and we propose a strategy for the detection of multiple waterborne parasites, including Cryptosporidium spp. and Giardia. Water samples were filtered to recover Toxoplasma oocysts and, after the detection of Cryptosporidium oocysts and Giardia cysts by immunofluorescence, as recommended by French norm procedure NF T 90-455, the samples were purified on a sucrose density gradient. Detection of Toxoplasma was based on PCR amplification and mouse inoculation to determine the presence and infectivity of recovered oocysts. After experimental seeding assays, we determined that the PCR assay was more sensitive than the bioassay. This strategy was then applied to 482 environmental water samples collected since 2001. We detected Toxoplasma DNA in 37 environmental samples (7.7%, including public drinking water; however, none of them were positive by bioassay. This strategy efficiently detects Toxoplasma oocysts in water and may be suitable as a public health sentinel method. Alternative methods can be used in conjunction with this one to determine the infectivity of parasites that were detected by molecular methods.
Feng, Yaoyu; Zhao, Xukun; Chen, Jiaxu; Jin, Wei; Zhou, Xiaonong; Li, Na; Wang, Lin; Xiao, Lihua
Genotyping studies on the source and human infection potential of Cryptosporidium oocysts in water have been almost exclusively conducted in industrialized nations. In this study, 50 source water samples and 30 tap water samples were collected in Shanghai, China, and analyzed by the U.S. Environmental Protection Agency (EPA) Method 1623. To find a cost-effective method to replace the filtration procedure, the water samples were also concentrated by calcium carbonate flocculation (CCF). Of the 50 source water samples, 32% were positive for Cryptosporidium and 18% for Giardia by Method 1623, whereas 22% were positive for Cryptosporidium and 10% for Giardia by microscopy of CCF concentrates. When CCF was combined with PCR for detection, the occurrence of Cryptosporidium (28%) was similar to that obtained by Method 1623. Genotyping of Cryptosporidium in 17 water samples identified the presence of C. andersoni in 14 water samples, C. suis in 7 water samples, C. baileyi in 2 water samples, C. meleagridis in 1 water sample, and C. hominis in 1 water sample. Therefore, farm animals, especially cattle and pigs, were the major sources of water contamination in Shanghai source water, and most oocysts found in source water in the area were not infectious to humans. Cryptosporidium oocysts were found in 2 of 30 tap water samples. The combined use of CCF for concentration and PCR for detection and genotyping provides a less expensive alternative to filtration and fluorescence microscopy for accurate assessment of Cryptosporidium contamination in water, although the results from this method are semiquantitative. PMID:21498768
or the synthetic peptide-based immunoassay (PeptiLav I and II), both obtained from Sanofi Diagnostic Pasteur, Marnes-la-. Coquette, France. Definition of diarrhoea episodes: The World Health. Organisation (WHO) criteria was used to determine diarrhoeal episodes (14). A total of three or more unformed stools in one 24-.
Giacometti, Andrea; Cirioni, Oscar; Del Prete, Maria Simona; Skerlavaj, Barbara; Circo, Raffaella; Zanetti, Margherita; Scalise, Giorgio
Two laboratory methods, a cell culture system and double fluorogenic staining, were used to study the viability and infective ability of Cryptosporidium parvum sporozoites and oocysts after short-term exposure to four cathelicidin peptides. The compounds, SMAP-29, BMAP-28, PG-1 and Bac7(1-35), exerted a strong cytotoxic effect on sporozoites, but did not affect the viability and function of oocysts consistently. Overall, in the sporozoite series, a percentage of the viable population decreased rapidly to less than detectable levels after 15 and 60 min exposure to the peptides at concentrations of 100 and 10 micro g/mL, respectively. In the oocyst series, no compound produced complete inhibition of parasite growth: 60-85% of the oocyst population was viable after 180 min exposure at 100 micro g/mL. SMAP-29 exerted the highest activity against both sporozoites and oocysts.
Jothikumar, N; da Silva, A J; Moura, I; Qvarnstrom, Y; Hill, V R
Rapid identification of the two major species of Cryptosporidium associated with human infections, Cryptosporidium hominis and Cryptosporidium parvum, is important for investigating outbreaks of cryptosporidiosis. This study reports the development and validation of a real-time PCR TaqMan procedure for detection of Cryptosporidium species and identification of C. hominis and C. parvum in stool specimens. This procedure comprised a generic TaqMan assay targeting the 18S rRNA for sensitive detection of Cryptosporidium species, as well as two other TaqMan assays for identification of C. hominis and C. parvum. The generic Cryptosporidium species assay can be duplexed with the C. parvum-specific assay. The generic Cryptosporidium species assay was able to detect ten Cryptosporidium species and did not cross-react with a panel of ten other protozoan parasites. The generic Cryptosporidium species assay could detect 1-10 oocysts in a 300 microl stool specimen, whilst each of the species-specific TaqMan assays had detection sensitivities that were approximately tenfold higher. The 18S rRNA assay was found to detect Cryptosporidium species in 49/55 DNA extracts from stool specimens containing either C. hominis or C. parvum. The C. hominis TaqMan assay correctly identified C. hominis in 24/31 validation panel specimens containing this species. The C. parvum-specific assay correctly identified C. parvum in 21/24 validation panel specimens containing this species. This real-time PCR procedure was used to detect and identify C. hominis and C. parvum in stool specimens from outbreak investigations in the USA and Botswana, resulting in identification of C. hominis and/or C. parvum in 66/67 stool specimens shown to be positive for these species using other techniques. From the outbreak specimens tested, the TaqMan procedure was found to have a specificity of 94%. This TaqMan PCR procedure should be a valuable tool for the laboratory diagnosis of cryptosporidiosis caused by C
Lee, Seung-Hyun; Joung, Migyo; Yoon, Sejoung; Choi, Kyoungjin; Park, Woo-Yoon; Yu, Jae-Ran
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.
Castro-Hermida, José Antonio; García-Presedo, Ignacio; Almeida, André; González-Warleta, Marta; Correia Da Costa, José Manuel; Mezo, Mercedes
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.
José Rafael Módolo
Full Text Available Foram examinadas, para pesquisa de Cryptosporidium pelo método de Heine,fezes de nove bezerros com criptosporidíase, após utilização prévia de dois diferentes desinfetantes. Quanto ao formol a 10%, notou-se que não houve interferência na identificação dos oocistos, em período compreendido entre cinco minutos e 72 horas; ao ser usado o hipoclorito de sódio a 14,5%, verificou-se que depois de 30minutos os ooçistos apresentaram-se avermelhados e sem refração, dificultando o reconhecimento. Assim, recomenda-se a adição de formol a 10% à matéria fecal, conforme a etapa referida, para coibir o risco de infecção de laboratoristas pelo vírus da imunodeficiência humana (HIV, quando usada para diagnóstico atécnica mencionada.Cryptosporidium oocysts were searched by Heine 's method in stools of nine cal/s with cryptoporidiosis after stool treatment with two disinfectants, 10% paraformaldehyde solution and 14,5% sodium hypochlorite solution. After 30 minutes exposition to sodium hypochlorite solution oocysts became non refractile and acquired a reddish tinge, making their Identification difficult. No morphological alterations occured in oocysts after paraformaldehyde treatment. We recommend paraformaldehyde at 10% concentration as means of human immunodeficiency virus (HIV inactivation for routine use in stool examinations and therefore making safer those type of procedures for laboratory personnel, when using Heine 's method.
Connelly, L.; Craig, B. H.; Jones, B.
This is the first report to characterize the genotypes and subtypes of Cryptosporidium species infecting a geographically isolated population of feral Soay sheep (Ovis aries) on Hirta, St. Kilda, Scotland, during two distinct periods: (i) prior to a population crash and (ii) as host numbers increased. Cryptosporidium DNA was extracted by freeze-thawing of immunomagnetically separated (IMS) bead-oocyst complexes, and species were identified following nested-PCR-restriction fragment length polymorphism (RFLP)/PCR sequencing at two Cryptosporidium 18S rRNA loci. Two hundred fifty-five samples were analyzed, and the prevalent Cryptosporidium species in single infections were identified as C. hominis (11.4% of all samples tested), C. parvum (9%), C. xiaoi (12.5%), and C. ubiquitum (6.7%). Cryptosporidium parvum was also present with other Cryptosporidium species in 27.1% of all samples tested. Cryptosporidium parvum- and C. hominis-positive isolates were genotyped using two nested-PCR assays that amplify the Cryptosporidium glycoprotein 60 gene (GP60). GP60 gene analysis showed the presence of two Cryptosporidium genotypes, namely, C. parvum IIaA19G1R1 and C. hominis IbA10G2. This study reveals a higher diversity of Cryptosporidium species/genotypes than was previously expected. We suggest reasons for the high diversity of Cryptosporidium parasites within this isolated population and discuss the implications for our understanding of cryptosporidiosis. PMID:23354707
Mary, C; Chapey, E; Dutoit, E; Guyot, K; Hasseine, L; Jeddi, F; Menotti, J; Paraud, C; Pomares, C; Rabodonirina, M; Rieux, A; Derouin, F
Cryptosporidium is a protozoan parasite responsible for gastroenteritis, especially in immunocompromised patients. Laboratory diagnosis of cryptosporidiosis relies on microscopy, antigen detection, and nucleic acid detection and analysis. Among the numerous molecular targets available, the 18S rRNA gene displays the best sensitivity and sequence variations between species and can be used for molecular typing assays. This paper presents a new real-time PCR assay for the detection and quantification of all Cryptosporidium species associated with the identification of Cryptosporidium hominis and Cryptosporidium parvum. The sensitivity and specificity of this new PCR assay were assessed on a multicentric basis, using well-characterized Cryptosporidium-positive and -negative human stool samples, and the efficiencies of nine extraction methods were comparatively assessed using Cryptosporidium-seeded stool samples and phosphate-buffered saline samples. A comparison of extraction yields showed that the most efficient extraction method was the Boom technique in association with mechanical grinding, and column extraction showed higher binding capacity than extraction methods based on magnetic silica. Our PCR assay was able to quantify at least 300 oocysts per gram of stool. Satisfactory reproducibility between laboratories was observed. The two main species causing human disease, Cryptosporidium hominis and Cryptosporidium parvum, were identified using a duplex real-time PCR assay with specific TaqMan minor-groove-binding ligand (MGB) probes for the same amplicon. To conclude, this one-step quantitative PCR is well suited to the routine diagnosis of cryptosporidiosis since practical conditions, including DNA extraction, quantification using well-defined standards, and identification of the two main species infecting humans, have been positively assessed.
Karasawa Andréa Satie Matsubara
Full Text Available The objective of the present study was to investigate the prevalence of Cryptosporidium (Apicomplexa, Cryptosporidiidae in the snake Crotalus durissus terrificus (Serpentes, Viperidae. Fifty animals were evaluated for the presence of oocysts of Cryptosporidium sp. at the time of arrival and 30 and 60 days later. Intestinal washings with saline solution (1% body weight, fecal samples, and organ scrapings were collected during the study. Oocysts were concentrated by an ether-phosphate-buffered saline sedimentation technique and then separated by a density gradient centrifugation technique. Smears were made with the sediment and submitted to modified acid-fast and auramine-rhodamine staining. Cryptosporidium-positive smears were used as controls for the experimental findings. The overall prevalence of Cryptosporidium sp. oocysts was 14%. Among the positive snakes, oocysts were detected only in the intestinal washing in two specimens, only in the feces in four specimens, and in both materials at least once in one specimen. The positive snakes were predominantly from Santa Maria da Serra city State of São Paulo (57.1%. We also observed that all of the examinations that presented positive results were obtained at least 27 days after the capture of the animals.
Petersen, H. H.; Wolsey, I.; Dalsgaard, A.
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...
Ranjbar-Bahadori, Sh; Mostoophi, A; Shemshadi, B
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.
Nakamura, Alex Akira; Simões, Daniel Castendo; Antunes, Rômulo Godik; da Silva, Deuvânia Carvalho; Meireles, Marcelo Vasconcelos
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).
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.
Sim, Seobo; Yu, Jae-Ran; Lee, Young-Ha; Lee, Jin-Su; Jeong, Hoo-Gn; Mohamed, Abd Al Wahab Saed; Hong, Sung-Tae
Cryptosporidium , a protozoan parasite that causes watery diarrhea, is found worldwide and is common in areas with low water hygiene. In February 2014, 866 stool samples were collected from the inhabitants of 2 rural areas in White Nile State, Sudan. These stool samples were assessed by performing modified acid-fast staining, followed by examination under a light microscope. The overall positive rate of Cryptosporidium oocysts was 13.3%. Cryptosporidium oocysts were detected in 8.6% stool samples obtained from inhabitants living in the area having water purification systems and in 14.6% stool samples obtained from inhabitants living in the area not having water purification systems. No significant difference was observed in the prevalence of Cryptosporidium infection between men and women (14.7% and 14.1%, respectively). The positive rate of oocysts by age was the highest among inhabitants in their 60s (40.0%). These findings suggest that the use of water purification systems is important for preventing Cryptosporidium infection among inhabitants of these rural areas in Sudan.
Li, Na; Neumann, Norman F.; Ruecker, Norma; Alderisio, Kerri A.; Sturbaum, Gregory D.; Villegas, Eric N.; Chalmers, Rachel; Monis, Paul; Feng, Yaoyu
The occurrence of Cryptosporidium oocysts in drinking source water can present a serious public health risk. To rapidly and effectively assess the source and human-infective potential of Cryptosporidium oocysts in water, sensitive detection and correct identification of oocysts to the species level (genotyping) are essential. In this study, we developed three real-time PCR genotyping assays, two targeting the small-subunit (SSU) rRNA gene (18S-LC1 and 18S-LC2 assays) and one targeting the 90-kDa heat shock protein (hsp90) gene (hsp90 assay), and evaluated the sensitivity and Cryptosporidium species detection range of these assays. Using fluorescence resonance energy transfer probes and melt curve analysis, the 18S-LC1 and hsp90 assays could differentiate common human-pathogenic species (C. parvum, C. hominis, and C. meleagridis), while the 18S-LC2 assay was able to differentiate nonpathogenic species (such as C. andersoni) from human-pathogenic ones commonly found in source water. In sensitivity evaluations, the 18S-LC2 and hsp90 genotyping assays could detect as few as 1 Cryptosporidium oocyst per sample. Thus, the 18S-LC2 and hsp90 genotyping assays might be used in environmental monitoring, whereas the 18S-LC1 genotyping assay could be useful for genotyping Cryptosporidium spp. in clinical specimens or wastewater samples. PMID:26092455
Matheus Hillard Farret; Vinicius da Rosa Fanfa; Luisa Ragagnin; Aleksandro Schafer da Silva; Silvia Gonzalez Monteiro
This study reports the gastrointestinal parasitism by Giardia sp. and Cryptosporidium sp. in blue macaw (Anodorhynchus hyacinthinus) in the southern region of Brazil. Fecal samples of two species kept in captivity in the state of Rio Grande do Sul were analyzed by the direct smear method, centrifugal flotation technique with zinc sulfate and Kinyoun staining technique for research of parasites. Mixed infection by eggs of Capillaria, cysts of Giardia sp. and oocysts of Cryptosporidium sp. was ...
Full Text Available Transport of Cryptosporidium in runoff will contaminate the surrounding water body. Cryptosporidium-sized microspheres and inactive Cryptosporidium parvum oocysts removal using an active carbon (AC filter with/without coal gangue from the runoff was evaluated. Chemical composition of coal gangue was determined, and its performance as a water treatment material was tested. Results showed SiO2 was the dominant chemical component of coal gangue. Removal was only 20% for microspheres and 24% for oocysts by AC filtration alone. The removal for both was increased to more than 98% by addition of coal gangue powder to the filter. Furthermore, gangue reuse turned commercially worthless material to a water treatment material as well helping prevent gangue from occupying agriculture/industry land.
Power, Michelle L; Ryan, Una M
Cryptosporidium macropodum n. sp is described. Oocysts of C. macropodum from the feces of kangaroos (Macropus spp.) are morphologically indistinguishable from other mammalian Cryptosporidium species, including C. parvum, C. hominis, C. suis, and C. canis. The oocysts are fully sporulated on excretion, lack sporocysts, and have an average width of 4.9 microm (4.5-6.0), a length of 5.4 microm (5.0-6.0), and a length:width ratio of 1.1. Phylogenetic analyses of the 18S ribosomal RNA, actin, and heat shock protein 70 (HSP70) loci demonstrate that C. macropodum is genetically distinct from all described Cryptosporidium species, including others found in marsupials. The parasite seems to be highly host-specific, because it has been found only in marsupials to date. Therefore, based on biological and molecular data, we consider C. macropodum a new species.
Full Text Available A cross-sectional survey was conducted to determine the prevalence of Cryptosporidium spp. infection in companion and stray dogs in Kerman, Iran. Faecal samples were randomly collected from 548 dogs (450 companion and 98 stray. Cryptosporidium oocysts were concentrated using the formalin ether sedimentation method according to the modified Ziehl-Neelsen staining technique. Cryptosporidium oocysts were identified in 2% (11/548 of samples. Faeces were classified according to the consistency as diarrhoeic (56/548 and non-diarrhoeic (492/548. Diarrhoea was recorded in 4 of the positive samples (7.14%. The prevalence of cryptosporidiosis was significantly higher in diarrhoeic dogs (7.14% compared to the non-diarrhoeic dogs (1.4% (p0.05. This study confirmed that dogs have a potential role in human cryptosporidiosis and faecal examination of dogs with persistent diarrhoea should be performed on a routine basis.
Vermeulen, Lucie; Hofstra, Nynke
Conclusions GloWPa-Crypto is the first global model that can be used to analyse dynamics in surface water pathogen concentrations worldwide. Global human Cryptosporidium emissions are estimated at 1 x 10^17 oocysts/ year for the year 2010.We estimated future emissions for SSP1 and SSP3. Preliminary results show that for SSP1human emissions are approximately halved by 2050. The SSP3 human emissions are 1.5 times higher than the 2010 emissions due to increased population growth and urbanisation. Livestock Cryptosporidium emissions are expected to increase under both SSP1 and SSP3, as meat consumption continues to rise. We conclude that population growth, urbanization, changes in sanitation systems and treatment, and changes in livestock consumption and production systems are important processes that determine future Cryptosporidium emissions to surface water. References Hofstra N, Bouwman A F, Beusen A H W and Medema G J 2013 Exploring global Cryptosporidium emissions to surface water Sci. Total Environ. 442 10-9 Kiulia N M, Hofstra N, Vermeulen L C, Obara M A, Medema G J and Rose J B 2015 Global occurrence and emission of rotaviruses to surface waters Pathogens 4 229-55 Vermeulen L C, De Kraker J, Hofstra N, Kroeze C and Medema G J 2015 Modelling the impact of sanitation, population and urbanization estimates on human emissions of Cryptosporidium to surface waters - a case study for Bangladesh and India Environ. Res. Lett. 10
Although standard methods for analyzing water samples for the protozoan parasites Cryptosporidium spp. and Giardia duodenalis are available and widely used, equivalent methods for analyzing water samples for Toxoplasma oocysts are lacking. This is partly due to the lack of a readily available, relia...
Lalancette, Cindy; Papineau, Isabelle; Payment, Pierre; Dorner, Sarah; Servais, Pierre; Barbeau, Benoit; Di Giovanni, George D; Prévost, Michèle
Assessing the presence of human pathogenic Cryptosporidium oocysts in surface water remains a significant water treatment and public health challenge. Most drinking water suppliers rely on fecal indicators, such as the well-established Escherichia coli (E. coli), to avoid costly Cryptosporidium assays. However, the use of E. coli has significant limitations in predicting the concentration, the removal and the transport of Cryptosporidium. This study presents a meta-analysis of E. coli to Cryptosporidium concentration paired ratios to compare their complex relationships in eight municipal wastewater sources, five agricultural fecal pollution sources and at 13 drinking water intakes (DWI) to a risk threshold based on US Environmental Protection Agency (USEPA) regulations. Ratios lower than the USEPA risk threshold suggested higher concentrations of oocysts in relation to E. coli concentrations, revealing an underestimed risk for Cryptosporidium based on E. coli measurements. In raw sewage (RS), high ratios proved E. coli (or fecal coliforms) concentrations were a conservative indicator of Cryptosporidium concentrations, which was also typically true for secondary treated wastewater (TWW). Removals of fecal indicator bacteria (FIB) and parasites were quantified in WWTPs and their differences are put forward as a plausible explanation of the sporadic ratio shift. Ratios measured from agricultural runoff surface water were typically lower than the USEPA risk threshold and within the range of risk misinterpretation. Indeed, heavy precipitation events in the agricultural watershed led to high oocyst concentrations but not to E. coli or enterococci concentrations. More importantly, ratios established in variously impacted DWI from 13 Canadian drinking water plants were found to be related to dominant fecal pollution sources, namely municipal sewage. In most cases, when DWIs were mainly influenced by municipal sewage, E. coli or fecal coliforms concentrations agreed with
Wang, Rongjun; Qi, Meng; Jingjing, Zhu; Sun, Dong; Ning, Changshen; Zhao, Jinfeng; Zhang, Longxian; Xiao, Lihua
Few data are available on the molecular characterization of Cryptosporidium spp. in ostriches. The objective of this study was to determine the prevalence of Cryptosporidium species or genotypes in ostriches. A total of 452 fecal samples from five farms, a zoo, and an animal rescue center in Zhengzhou, Henan Province, China were examined for Cryptosporidium oocysts by microscopy of wet mount of fecal materials concentrated by the Sheather's sugar flotation technique. Fifty-three samples were Cryptosporidium-positive from four farms, with an overall prevalence of 11.7%. The percentage of animals shedding oocysts was 0, 16.2%, 7.2%, and 0 in 1-3 weeks, 4-8 weeks, 3-12 months, and more than 12 months ostriches, respectively (χ(2)=17.74; ρ<0.01). PCR-restriction fragment length polymorphism (RFLP) analysis of the small subunit (SSU) rRNA gene of the 53 Cryptosporidium-positive samples showed the presence of only Cryptosporidium baileyi, which was confirmed by DNA sequencing of the SSU rRNA PCR products from 16 positive samples. Cross-transmission studies demonstrated that the C. baileyi isolate could infect chickens and quails. Thus, ostriches are commonly infected with C. baileyi that is genetically and biologically similar to C. baileyi found in other birds. Copyright © 2010 Elsevier B.V. All rights reserved.
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
Wang, Rongjun; Zhang, Longxian; Feng, Yaoyu; Ning, Changshen; Jian, Fuchun; Xiao, Lihua; Zhao, Jinfeng; Wang, Yongli
A total of 469 fecal samples were collected from American minks (Mustela vison) on a farm in Hebei Province in China and examined for Cryptosporidium by Sheather's sugar flotation technique and 8 Cryptosporidim isolates were obtained. The partial 18S rRNA, 70kDa heat shock protein (HSP70), Cryptosporidium oocyst wall protein (COWP) and actin genes of six isolates were sequenced. Sequence data were analyzed together with known Cryptosporidium spp. and genotypes. Results of this multi-locus genetic characterization indicated that the six Cryptosporidium isolates in this study shared the same sequences of the genes studied and were different from known Cryptosporidium species and genotypes. The closest relative was Cryptosporidium ferret genotype with 7, 22, 2 and 2 nucleotide differences in the 18S rRNA, HSP70, COWP and actin genes, respectively. The homology to ferret genotype at the 18S rRNA locus was 99.1%, which is comparable to that between C. parvum and C. hominis (99.2%), or between C. muris and C. andersoni (99.4%). Therefore, the Cryptosporidium in minks in this study is considered a new genotype, the Cryptosporidium mink genotype.
Soltane, R; Guyot, K; Dei-Cas, E; Ayadi, A
A longitudinal study was undertaken to determine the prevalence of Cryptosporidium in a dairy farm in Sfax, Tunisia. 480 faecal samples were obtained from 30 calves under one month of age. All faecal samples were analysed for Cryptosporidium oocysts by microscopic examination of smears stained by modified Ziehl Neelsen technique. The parasite was detected in 26 calves (86.7%). Infection was significantly associated with diarrhoea. A molecular characterization, performed in seven calves, confirmed that isolates were C. parvum. This work is the first report on Cryptosporidium in calves in Tunisia.
Marounek, Milan; Tůmová, E.; Skřivan, M.; Pavlásek, I.; Ledvinka, Z.
Roč. 46, - (2002), s. 203-207 ISSN 0005-2086 R&D Projects: GA ČR GA524/99/0480; GA AV ČR KSK5020115 Keywords : chickens * experimental cryptosporidiosis * performance Subject RIV: ED - Physiology Impact factor: 1.151, year: 2002
Moore, Catrin E; Elwin, Kristin; Phot, Nget; Seng, Chanthou; Mao, Saroeun; Suy, Kuong; Kumar, Varun; Nader, Johanna; Bousfield, Rachel; Perera, Sanuki; Bailey, J Wendi; Beeching, Nicholas J; Day, Nicholas P J; Parry, Christopher M; Chalmers, Rachel M
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 (pCryptosporidium 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 sanitation, increase hand washing after defecation and before preparing food and promote drinking boiled water may reduce the burden of these two parasites.
Catrin E Moore
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
Moore, Catrin E.; Elwin, Kristin; Phot, Nget; Seng, Chanthou; Mao, Saroeun; Suy, Kuong; Kumar, Varun; Nader, Johanna; Bousfield, Rachel; Perera, Sanuki; Bailey, J. Wendi; Beeching, Nicholas J.; Day, Nicholas P. J.; Parry, Christopher M.; Chalmers, Rachel M.
Background 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. Methodology/Principal Findings 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). Conclusions/Significance 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
Abd El Kader, Nour M; Blanco, María-Alejandra; Ali-Tammam, Marwa; Abd El Ghaffar, Abd El Rahman B; Osman, Ahmed; El Sheikh, Nabila; Rubio, José Miguel; de Fuentes, Isabel
Cryptosporidium is a significant cause of diarrheal disease in developing and industrialized nations. Cryptosporidium hominis and Cryptosporidium parvum are the main agents of cryptosporidiosis in humans. In Egypt, very little is known about genetic structure of Cryptosporidium spp. Therefore, this study was designed to examine samples from sporadic cases of cryptosporidiosis in Egyptians in order to identify the species involved in infection as well as the transmission dynamics and distribution of the parasite in the Great Cairo area. A total of 391 human faecal samples were collected, between May 2008 and March 2009, from ten public hospitals in Great Cairo. Initial screening by immunochromatographic detection kit "the Stick Crypto-Giardia; Operon" showed 23 possible positive cases. Twenty of them were confirmed by microscopic examination. PCR was performed by amplification of the oocyst wall protein (COWP) gene where 18 out of 23 samples were positive, one not detected by microscopy. Cryptosporidium genotyping was performed by RFLP analysis of PCR products of the diagnosis PCR. Only 15 samples rendered a digestion pattern. The genotyping distribution was nine cases showing C. hominis genotype, three showing C. parvum genotype and three showing mixed infection by C. hominis and C. parvum. The data showed an elevated prevalence of C. hominis (80.0%), the most anthroponotic species, suggesting a human-human transmission. Furthermore, the presence of up to 40% of samples infected with C. parvum shows that further investigations are required to determine the subgenotypes of C. parvum to clarify the mode of transmission in order to improve the control measures.
Maria Aparecida da Glória Faustino
Full Text Available The apicomplexa protozoa Cryptosporidium infects several mammals, including terrestrial and aquatic species. In the epidemiology of this infection, the ingestion of water and/or food contamined with oocysts comprises the main mechanism of transmission to susceptible animals. Among the Sirenians, the occurrence of this coccidium has been reported in dugongs (Dugong dugon and Antillean manatee (Trichechus manatus manatus. The present study was conducted with the aim of verifying the occurrence of Cryptosporidium spp. in Amazonian manatee. For this purpose, fecal samples were collected from ten free-ranging Amazonian manatees, two specimens in captivity, and 103 supernatants fecal samples. The samples were processed by the sedimentation method in formol-ether and Kinyoun stain technique for the presence of Cryptosporidium spp.. The positive samples were then submitted to Direct Immunoflorescence Test. The results showed 4.34% (05/115 of positive samples. This is the first report of Cryptosporidium spp. in the Amazonian manatee.
Monis, P T; Saint, C P
A nested-PCR assay, incorporating an internal positive control, was developed for Cryptosporidium monitoring in finished water. This assay was capable of reproducibly detecting 8 oocysts in spiked-filtered water samples collected from 5 South Australian water treatment plants. The RT-PCR assay of Kaucner and Stinear (Appl. Environ. Microbiol. 64(5) (1998) 1743) was also evaluated for the detection of Cryptosporidium parvum. Initially, under our experimental conditions, a detection level of 27 oocysts was achieved for spiked reagent water samples. This level was improved to 5 oocysts by modification of the method. Untreated South Australian source waters concentrated by calcium carbonate flocculation were found to be highly inhibitory to the RT-PCR assay. Concentration of similar samples using Envirochek filters appeared to eliminate PCR inhibition. While both methods possessed similar sensitivities the nested-PCR assay was more reproducible, more cost effective, simpler to perform and could detect both viable and non-viable intact Cryptosporidium parvum oocysts, which is an important consideration for plant operators. These factors make the nested-PCR assay the method of choice for screening large numbers of potable water samples, where a reliable low level of detection is essential.
Hijjawi, Nawal; Estcourt, Annika; Yang, Rongchang; Monis, Paul; Ryan, Una
The present study reports for the first time the completion of the life cycle of Cryptosporidium hominis in cell-free culture and multiplication of the parasite via qPCR. Individual life-cycle stages were characterised using Cryptosporidium-specific antibody staining (Sporo-Glo) and fluorescent in situ hybridisation (FISH) staining on cultures inoculated with excysted oocysts and purified sporozoites. In both cultures, C. hominis successfully proliferated and completed its life cycle, however development in cultures inoculated with purified sporozoites lagged behind cultures inoculated with excysted oocysts. Some novel findings of the study include the visualisation of pairing and multiple associations between various developmental stages in a process similar to syzygy and the formation of Cryptosporidium stages (trophozoites and meronts) inside the oocysts without excystation. qPCR analysis revealed a 5-6-fold amplification of parasite DNA. Future studies are required to improve the amplification of the parasite. The present study confirms the suitability of this culturing model to support the growth and proliferation of C. hominis (which unlike C. parvum, cannot be readily cultured in small animal models) and will greatly assist in our understanding of the developmental biology of Cryptosporidium, its position within the Apicomplexa and its relationship to gregarine protozoa.
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...
Wang, Hui-zhu; Jiao, Bing-xin; Tian, Jing-hua; Li, Min; Guo, Jie; Liu, Ying; Li, Xing-wang; Wang, Yu-guang
To investigate the Cryptosporidium infection and its epidemiological characteristics in HIV/AIDS patients with chronic diarrhea. Stool samples collected from HIV/AIDS confirmed patients with chronic diarrhea who lived in Beijing, Henan and Xinjiang. Samples were concentrated by Formalin-Ethyl Acetate Sedimentation technique and stained by modified acid-fast stain (AFS) for the identification of oocysts by microscopy. CD4(+)T cells count was performed by Flow Cytometry. The overall infection rate of Cryptosporidium in AIDS patients was 12.6% (32/253). The infection rates of oocysts in the area of Beijing, Henan and Xinjiang were 5.97% (4/67), 16.1% (24/149) and 10.8% (4/37) respectively. The infection rate of oocysts in the urban areas was 6.5% (7/104) while in the countryside it was 16.8% (25/149) and the difference was significantly different. However, there were no any differences discovered between the infection rates on patient's gender or on infection occurred in different seasons. The infectious rates of oocyst in patients on different stages of the disease were also significantly different (P AIDS patients infected by Cryptosporidium were not rarely seen in northern China. The rate of infection was not associated with patient's gender but was associated with patient's living environments. Patients living in the countryside, with lower lever of CD4(+)T cells counts and at the middle/late stage of the disease, Cryptosporidium infection appeared to be high.
Guo, Yaqiong; Li, Na; Lysén, Colleen; Frace, Michael; Tang, Kevin; Sammons, Scott; Roellig, Dawn M; Feng, Yaoyu; Xiao, Lihua
Whole-genome sequencing of Cryptosporidium spp. is hampered by difficulties in obtaining sufficient, highly pure genomic DNA from clinical specimens. In this study, we developed procedures for the isolation and enrichment of Cryptosporidium genomic DNA from fecal specimens and verification of DNA purity for whole-genome sequencing. The isolation and enrichment of genomic DNA were achieved by a combination of three oocyst purification steps and whole-genome amplification (WGA) of DNA from purified oocysts. Quantitative PCR (qPCR) analysis of WGA products was used as an initial quality assessment of amplified genomic DNA. The purity of WGA products was assessed by Sanger sequencing of cloned products. Next-generation sequencing tools were used in final evaluations of genome coverage and of the extent of contamination. Altogether, 24 fecal specimens of Cryptosporidium parvum, C. hominis, C. andersoni, C. ubiquitum, C. tyzzeri, and Cryptosporidium chipmunk genotype I were processed with the procedures. As expected, WGA products with low (sequences in Sanger sequencing. The cloning-sequencing analysis, however, showed significant contamination in 5 WGA products (proportion of positive colonies derived from Cryptosporidium genomic DNA, ≤25%). Following this strategy, 20 WGA products from six Cryptosporidium species or genotypes with low (mostly sequencing, generating sequence data covering 94.5% to 99.7% of Cryptosporidium genomes, with mostly minor contamination from bacterial, fungal, and host DNA. These results suggest that the described strategy can be used effectively for the isolation and enrichment of Cryptosporidium DNA from fecal specimens for whole-genome sequencing. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
Miller, Christopher N; Jossé, Lyne; Brown, Ian; Blakeman, Ben; Povey, Jane; Yiangou, Lyto; Price, Mark; Cinatl, Jindrich; Xue, Wei-Feng; Michaelis, Martin; Tsaousis, Anastasios D
Cryptosporidium parasites are a major cause of diarrhoea that pose a particular threat to children in developing areas and immunocompromised individuals. Curative therapies and vaccines are lacking, mainly due to lack of a long-term culturing system of this parasite. Here, we show that COLO-680N cells infected with two different Cryptosporidium parvum strains produce sufficient infectious oocysts to infect subsequent cultures, showing a substantial fold increase in production, depending on the experiment, over the most optimistic HCT-8 models. Oocyst identity was confirmed using a variety of microscopic- and molecular-based methods. This culturing system will accelerate research on Cryptosporidium and the development of anti-Cryptosporidium drugs. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.
Hadfield, Stephen J.; Robinson, Guy; Elwin, Kristin; Chalmers, Rachel M.
Real-time PCR has the potential to streamline detection and identification of Cryptosporidium spp. in human clinical samples. In the present article, we report the first such assay to allow not only detection and differentiation of the most common human pathogens, Cryptosporidium hominis and Cryptosporidium parvum, but also simultaneous amplification of a region of the small subunit (SSU) rRNA gene, permitting direct sequence analysis to identify any Cryptosporidium species. An internal control is incorporated to identify the presence of PCR inhibitors. Analytical sensitivity was determined to be as low as 200 oocysts per gram of feces processed, equivalent to 2 oocysts per PCR. The C. hominis and C. parvum PCRs specifically detected only species/genotypes in their respective target clades. Diagnostic sensitivity and specificity, evaluated against a widely used conventional nested SSU rRNA gene PCR as a nominated gold standard using a panel of 258 (151 positive and 107 negative) samples, were 100% and 99.1%, respectively. The assay agreed with PCR-restriction fragment length polymorphism analysis of the Cryptosporidium oocyst wall protein gene for 134 of 136 (98.5%) samples tested prospectively and typed two additional isolates. The real-time PCR assay was sensitive, specific, and reproducible and significantly improved laboratory work flow and turnaround times. PMID:21177904
Deng, Lei; Li, Wei; Zhong, Zhijun; Gong, Chao; Cao, Xuefeng; Song, Yuan; Wang, Wuyou; Huang, Xiangming; Liu, Xuehan; Hu, Yanchun; Fu, Hualin; He, Min; Wang, Ya; Zhang, Yue; Wu, Kongju; Peng, Guangneng
A total of 333 fecal specimens from horses in southwestern China were genotyped based on analysis of the small subunit rRNA (SSU rRNA) gene. Cryptosporidium hominis and Cryptosporidium andersoni were identified in 2 and 4 stool specimens, respectively. The identification of C. hominis was confirmed by sequence analysis of the 70-kDa heat shock protein (HSP70) and oocyst wall protein (COWP) genes. Subtyping analysis of the 60-kDa glycoprotein (GP60) gene sequence of C. hominis revealed a new rare subtype Id, named IdA15; only three Id isolates have been reported in humans to date. Multilocus sequence typing (MLST) analysis indicated that the C. andersoni subtype was A6, A5, A2, and A1 at the four minisatellite loci (MS1, MS2, MS3, and MS16, respectively). This is the first report to identify the presence of C. andersoni and C. hominis in horses in southwestern China and the first to identify a rare zoonotic subtype Id of C. hominis in horses. These findings suggest that infected horses may act as potential reservoirs of Cryptosporidium to transmit infections to humans. © 2017 The Authors Journal of Eukaryotic Microbiology published by Wiley Periodicals, Inc. on behalf of International Society of Protistologists.
Cryptosporidium Zoonosis in Nigeria. Ayinmode, A. B. and Fagbemi B. O. Department of Veterinary Microbiology and Parasitology,. Faculty of Veterinary Medicine, University of Ibadan, Ibadan, Nigeria. ABSTRACT: Cryptosporidium is a coocidian parasite that infects a wide range of vertebrate hosts including man.
Vandenberg, Olivier; Robberecht, Françoise; Dauby, Nicolas; Moens, Catherine; Talabani, Hana; Dupont, Eddy; Menotti, Jean; van Gool, Tom; Levy, Jack
Cryptosporidium outbreaks in day-care centers (DCCs) occur commonly. However, controlling spread of infection in these settings is difficult, and data about effectiveness of different control strategies are sparse. In this study, a Cryptosporidium outbreak in a large DCC located in Brussels is described with evaluation of hygienic and therapeutic interventions. During a 3-week period, 43 of 130 children attending the DCC developed enteric symptoms. Stools from 122 children were examined for microbial pathogens. Of them, 38 (31%) were diagnosed with Cryptosporidium, 29 of them being symptomatic (76%) and 9 (24%) asymptomatic. Diagnosis was performed by microscopy, antigen tests, and real-time polymerase chain reaction. Strict infection control measures were implemented during the first week after the start of outbreak. After 4 weeks, 27/38 children (71%) were still symptomatic and Cryptosporidium positive. Because of persisting symptoms and fear of further spread of infection, all 27 children were treated with paromomycin. Two weeks later, 18 of 27 children were asymptomatic and were parasitologically negative. The remaining 9 children, still symptomatic and Cryptosporidium positive, were treated with nitazoxanide. Three weeks later, week 9 after the start of outbreak, all 38 children involved in the outbreak were asymptomatic and Cryptosporidium negative. Our study underscores the need to rule out Cryptosporidium etiology in a diarrheal outbreak in a DCC. Rapid implementation of infection control measures can most likely halt the spread of infection. The role of nitazoxanide to limit duration of shedding of oocysts deserves more attention for its use in outbreaks.
Liu, Aiqin; Zhang, Jia; Zhao, Jingmin; Zhao, Wei; Wang, Rongjun; Zhang, Longxian
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.
Lv, Chaochao; Zhang, Longxian; Wang, Rongjun; Jian, Fuchun; Zhang, Sumei; Ning, Changshen; Wang, Helei; Feng, Chao; Wang, Xinwei; Ren, Xupeng; Qi, Meng; Xiao, Lihua
To understand the prevalence of Cryptosporidium infection in rodents in China and to assess the potential role of rodents as a source for human cryptosporidiosis, 723 specimens from 18 rodent species were collected from four provinces of China and examined between August 2007 and December 2008 by microscopy after using Sheather's sugar flotation and modified acid-fast staining. Cryptosporidium oocysts were detected in 83 specimens, with an overall prevalence of 11.5%. Phodopus sungorus, Phodopus campbelli, and Rattus tanezumi were new reported hosts of Cryptosporidium. The genotypes and subtypes of Cryptosporidium strains in microscopy-positive specimens were further identified by PCR and sequence analysis of the small subunit rRNA and the 60-kDa glycoprotein (gp60) genes. In addition to Cryptosporidium parvum, C. muris, C. andersoni, C. wrairi, ferret genotype, and mouse genotype I, four new Cryptosporidium genotypes were identified, including the hamster genotype, chipmunk genotype III, and rat genotypes II and III. Mixed Cryptosporidium species/genotypes were found in 10.8% of Cryptosporidium-positive specimens. Sequence analysis of the gp60 gene showed that C. parvum strains in pet Siberian chipmunks and hamsters were all of the subtype IIdA15G1, which was found previously in a human isolate in The Netherlands and lambs in Spain. The gp60 sequences of C. wrairi and the Cryptosporidium ferret genotype and mouse genotype I were also obtained. These findings suggest that pet rodents may be potential reservoirs of zoonotic Cryptosporidium species and subtypes. PMID:19820152
Oliveira, Bruno César Miranda; Ferrari, Elis Domingos; da Cruz Panegossi, Mariele Fernanda; Nakamura, Alex Akira; Corbucci, Flávio Sader; Nagata, Walter Bertequini; Dos Santos, Bianca Martins; Gomes, Jancarlo Ferreira; Meireles, Marcelo Vasconcelos; Widmer, Giovanni; Bresciani, Katia Denise Saraiva
The carrier pigeon and the domestic pigeon are different breeds of the species Columba livia. Carrier pigeons are used for recreational activities such as bird contests and exhibitions. Due to the close contact with humans, these birds may potentially represent a public health risk, since they can host and disseminate zoonotic parasites, such as those belonging to the genus Cryptosporidium (phylum Apicomplexa). The purpose of this work was the detection by microscopic and molecular techniques of Cryptosporidium spp. oocysts in fecal samples of carrier pigeons, and subsequently to sequence the 18S ribosomal RNA marker of positive samples to identify the species. A total of 100 fecal samples were collected individually in two pigeon breeding facilities from Formiga and Araçatuba, cities located in Minas Gerais state and São Paulo state, Brazil, respectively. The age of the birds ranged from one to 12 years; 56 were females and 44 males. Fecal smears were stained with negative malachite green, whereas the molecular characterization was based on the sequence of a ∼800bp fragment of the 18S rRNA gene. Microscopic examination of fecal smears revealed 4% (4/100) oocyst positivity. On the other hand, 7% (7/100) of positivity were found using nested PCR. Three samples were 99% to 100% similar to Cryptosporidium parvum 18S rDNA type A (Genbank AH006572) and the other three samples had 99% to 100% similarity to C. parvum 18S rDNA type B (Genbank AF308600). To our knowledge, this is the first report of C. parvum oocysts in carrier pigeons. Copyright © 2017 Elsevier B.V. All rights reserved.
Fontán-Sainz, María; Gómez-Couso, Hipólito; Fernández-Ibáñez, Pilar; Ares-Mazás, Elvira
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).
Ferrari, Elis Domingos; Nakamura, Alex Akira; Nardi, Ana Rita Moraes; Santana, Bruna Nicoleti; da Silva Camargo, Vinicius; Nagata, Walter Bertequini; Bresciani, Katia Denise Saraiva; Meireles, Marcelo Vasconcelos
The aim of this study was to evaluate the prevalence of and diagnostic methods for Cryptosporidium spp. in caged adult exotic parrots from Southern and Southeastern Brazil. Oocysts were purified from fecal samples from 463 psittacines by centrifugal-flotation in Sheather's sugar solution. Cryptosporidium spp. were detected by malachite green negative staining and nested PCR targeting the 18S rRNA gene. Cryptosporidium species were identified by sequencing nested PCR amplicons. Samples were also tested by duplex real-time PCR targeting the 18S rRNA gene of Cryptosporidium galli and Cryptosporidium avian genotype III. The prevalence rates of Cryptosporidium spp. determined by microscopy and nested PCR were 3.0% (14/463) and 5.0% (23/463), respectively. The nested PCR/sequencing identified avian genotype III (1.7%; 8/463), Cryptosporidium parvum (0.9%; 4/463) and Cryptosporidium canis (0.2%; 1/463). Duplex real-time PCR was positive for gastric Cryptosporidium in 9.5% (44/463) of the samples. Among them, 1.9% (9/463) were positive for C. galli, 5.8% (27/463) were positive for avian genotype III and 1.7% (8/463) showed mixed infections with C. galli and avian genotype III. With regards to the positive detection of Cryptosporidium spp., there was no statistically significant difference between nested PCR and microscopic analysis (p = .1237), and a fair agreement existed between them (Kappa = 0.242). A statistically significant difference (p < .0001) and fair agreement (Kappa = 0.317) were obtained between nested PCR/sequencing and duplex real-time PCR for the detection of gastric Cryptosporidium. We determined that nested PCR and duplex real-time PCR are the best options for the detection of Cryptosporidium spp. and gastric Cryptosporidium, respectively, and that avian genotype III is the most common Cryptosporidium genotype/species in psittacines. Copyright © 2017 Elsevier Inc. All rights reserved.
Sáez-Llorens, X; Odio, C M; Umaña, M A; Morales, M V
The efficacy of spiramycin was evaluated in a double blind, placebo-controlled study of 44 immunocompetent infants ages 2 to 13 months who had acute diarrhea caused by Cryptosporidium. Twenty-one patients received spiramycin (100 mg/kg/day) for 10 days and 23 received placebo. On admission the patients in both groups were comparable regarding demographic and clinical characteristics. The infants who were treated with spiramycin had a shorter duration of diarrhea (mean, 5.2 vs. 7.3 days; P = 0.002) and a shorter duration of excretion of oocysts in the stools (7.1 vs. 8.5 days; P = 0.032) compared with those treated with placebo. No clinical or parasitologic relapses were seen in patients of both groups. Mild adverse effects to spiramycin were observed in 2 patients (10%). Spiramycin appeared to hasten clinical recovery and decrease the duration of oocyst excretion in immunocompetent children with diarrheal illness caused by Cryptosporidium.
Moreira, Maria João; Soares, Sónia; de Lurdes Delgado, Maria; Figueiredo, João; Magalhães, Elisabete Silva; Castro, António; Viana Da Costa, Alexandra; Correia da Costa, José Manuel
To understand the situation of water contamination with Cryptosporidium spp. and Giardia spp. in the northern region of Portugal, we have established a long-term program aimed at pinpointing the sources of surface water and environmental contamination, working with the water-supply industry. Here, we describe the results obtained with raw water samples collected in rivers of the 5 hydrographical basins. A total of 283 samples were analyzed using the Method 1623 EPA, USA. Genetic characterization was performed by PCR and sequencing of genes 18S rRNA of Cryptosporidium spp. and β-giardin of Giardia spp. Infectious stages of the protozoa were detected in 72.8% (206 of 283) of the water samples, with 15.2% (43 of 283) positive for Giardia duodenalis cysts, 9.5% (27 of 283) positive for Cryptosporidium spp. oocysts, and 48.1% (136 of 283) samples positive for both parasites. The most common zoonotic species found were G. duodenalis assemblages A-I, A-II, B, and E genotypes, and Cryptosporidium parvum, Cryptosporidium andersoni, Cryptosporidium hominis, and Cryptosporidium muris. These results suggest that cryptosporidiosis and giardiasis are important public health issues in northern Portugal. To the authors' knowledge, this is the first report evaluating the concentration of environmental stages of Cryptosporidium and Giardia in raw water samples in the northern region of Portugal. PMID:20585525
Kváč, Martin; Hofmannová, L.; Bertolino, S.; Wauters, L.; Tosi, G.; Modrý, David
Roč. 55, č. 2 (2008), s. 95-99 ISSN 0015-5683 R&D Projects: GA ČR GP523/07/P117; GA ČR GA524/05/0992 Institutional research plan: CEZ:AV0Z60220518 Keywords : Cryptosporidium * Sciurus vulgaris * 18S rRNA * oocyst morphology * infectivity * red squirrel Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 1.307, year: 2008
Balatbat, A B; Jordan, G W; Tang, Y J; Silva, J
Cryptosporidium parvum is a coccidian protozoan that causes diarrhea in humans, often chronic and severe in patients with AIDS. Conventionally, diagnosis is made by concentration of stools followed by acid-fast staining (AF) or immunofluorescent staining. The threshold of detection in human stool specimens by these methods may require the presence of 50,000 (immunofluorescent staining) to 500,000 (AF) oocysts per g of stool. In this study, a nested PCR assay was developed to detect C. parvum ...
Amimul M Ehsan
Full Text Available 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
Taha, Shahinaz; Elmalik, Khitma; Bangoura, Berit; Lendner, Matthias; Mossaad, Ehab; Daugschies, Arwid
Cryptosporidiosis is a common protozoan infection causing morbidity and mortality in young cattle and may be zoonotically transmitted to humans. So far, there is no data available on the presence of Cryptosporidium spp. in the Sudan. The aim of this study was to isolate, identify, and genotype Cryptosporidium oocysts sampled from diarrheic calves housed at different farms in three different municipalities in Khartoum State (Khartoum, Khartoum North, Omdurman). A total of 149 fecal samples were evaluated microscopically for the presence of Cryptosporidium oocysts using the modified Ziehl-Neelsen staining method and 87 (58.3%) samples tested positive. Positive and negative samples were further analyzed by nested PCR targeting the SSU rRNA region. Positive samples were subjected to restriction enzyme analysis of PCR amplicons (PCR-RFLP). Nested PCR identified Cryptosporidium DNA in 53 samples (35.5%); restriction digestion of the PCR products revealed the presence of C. parvum (73.5%), C. ryanae (13.2%), C. andersoni (7.5%), and C. bovis (1.8%). Species distribution was clearly related to age with C. parvum being the predominant species in dysenteric pre-weaned calves. Sequencing of three genes (SSU rRNA, COWP, and GP60) for three C. parvum isolates originating from the three different municipalities showed that all belong to C. parvum subtype family IId. Based on data obtained by GP60, sequencing the two C. parvum isolates from Khartoum and Omdurman represent subtype IIdA18G1, whereas oocysts isolated in Khartoum North belong to subtype IIdA19G1. The observed genotypes are zoonotic and thus C. parvum in calves is potentially a health risk to humans in Khartoum State, Sudan. To the best of our knowledge, this is the first reported attempt to characterize Cryptosporidium isolated from cattle in the Sudan.
Siwila, J.; Phiri, I. G. K.; Enemark, Heidi L.
Cryptosporidium spp. and Giardia duodenalis are important parasites infecting a wide range of domestic animals worldwide. The aim of the present study was to determine the occurrence of Cryptosporidium spp. and Giardia parasites in different domestic animals living in close contact with humans...... within rural/semiurban communities in Kafue district in Zambia. A single faecal sample per animal was collected from pigs, goats, dogs, ducks, chickens and pigeons and analysed by Merifluor Cryptosporidium/Giardia immunofluorescence antibody assay for the simultaneous detection of these parasites....... The faecal consistency was noted and scored as non-diarrhoeic or diarrhoeic. A total of 236 samples were collected. Cryptosporidium spp. oocysts were detected in pigs (11.5%, 17/148), goats (5.9%; 1/17), ducks (10.0%; 3/30) and chickens (14.3%; 2/14) while Giardia cysts were detected in pigs (8.1%; 12...
Bouzid, Maha; Chalmers, Rachel M.; Tyler, Kevin M.
Cryptosporidium is a protozoan parasite of medical and veterinary importance that causes gastroenteritis in a variety of vertebrate hosts. Several studies have reported different degrees of pathogenicity and virulence among Cryptosporidium species and isolates of the same species as well as evidence of variation in host susceptibility to infection. The identification and validation of Cryptosporidium virulence factors have been hindered by the renowned difficulties pertaining to the in vitro culture and genetic manipulation of this parasite. Nevertheless, substantial progress has been made in identifying putative virulence factors for Cryptosporidium. This progress has been accelerated since the publication of the Cryptosporidium parvum and C. hominis genomes, with the characterization of over 25 putative virulence factors identified by using a variety of immunological and molecular techniques and which are proposed to be involved in aspects of host-pathogen interactions from adhesion and locomotion to invasion and proliferation. Progress has also been made in the contribution of host factors that are associated with variations in both the severity and risk of infection. Here we provide a review comprised of the current state of knowledge on Cryptosporidium infectivity, pathogenesis, and transmissibility in light of our contemporary understanding of microbial virulence. PMID:23297262
Full Text Available Cryptosporidium and Giardia are protozoan parasites transmitted by fecal-oral ingestion of (oocysts, and are responsible for enteritis in several animal species and humans worldwide. These (oocysts can survive for over a year in aquatic environments and can accumulate in bivalve mollusks, which filter large volumes of water. The aim of this study is to evaluate the natural occurrence of Cryptosporidium and Giardia contamination in different specimens of edible bivalves mollusks from farming sites of the western and north-eastern coasts of Sardinia. From April 2011 to February 2012, 1095 specimens of Mytilus galloprovincialis and 240 of Crassostrea gigas were sampled from Olbia and Oristano gulf and San Teodoro pond. Hepatopancreas and gills, including the labial palp, were examined for oocysts and cysts after pooling and homogenisation using different techniques: i staining for light and fluorescence microscopy; ii direct immunofluorescence (IF Merifluor® test Cryptosporidium/ Giardia (Meridian Bioscience Inc., Cincinnati, OH, USA; and iii molecular procedures. However, in the context under study, all mollusks examined with the three main diagnostic techniques were negative for both parasites pointing out the hypothetically low zoonotic risk related to Cryptosporidium and Giardia in bivalves, especially Mytilus galloprovincialis and Crassostrea gigas.
Zhao, Z; Wang, R; Zhao, W; Qi, M; Zhao, J; Zhang, L; Li, J; Liu, A
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.
Anejo-Okopi, Joseph Aje; Okojokwu, Julius Ocheme; Ebonyi, Augustine Odo; Ejeliogu, Emeka Uba; Isa, Samson Ejiji; Audu, Onyemocho; Akpakpan, Edoama Edet; Nwachukwu, Esther Ebere; Ifokwe, Christabel Kelechi; Ali, Murna; Lar, Patricia; Oguche, Stephen
Introduction Cryptosporidium is an important cause of diarrhea in children and immune-compromised individuals. Recent advances in molecular diagnostics have led to the discovery of subtype families that are thought to be more commonly associated with diarrhea. We aimed to isolate and characterize Cryptosporidium spp among children with diarrhea in Jos, Nigeria. Methods Stool samples were collected from165 children aged 0-5 years with diarrhea. Cryptosporidium oocysts were examined by wet mount preparation, using formalin ether and a modified acid fast staining method. DNA was extracted from positive samples using QIAamp DNA stool mini kit and PCR-RFLP assay was carried out after quantification. Genotyping and phylogenetic analysis were done to determine the subtype families and their relatedness. Results From the 165 children studied, 8 (4.8%) were infected with Cryptosporidium. PCR-RFLP assay and genotype characterization found the following Cryptosporidium species: C. hominis 6 (75%) and C. parvum 2 (25.0%), with family subtypes Id-5, Ie-1 and IIa-1, IId-1 respectively.The most common species was C. hominis and the frequent subtype was C. hominis-Id 5 (62.5%). Conclusion Cryptosporidium is not an uncommon cause of diarrhea in children, with C. hominis being the dominant species. Also C. hominis Id is the commonest sub-family subtype. Put together, zoonotic species may be an important cause of diarrhea in children aged 0-5 years in Jos, Nigeria. PMID:28293369
Reboredo-Fernández, Aurora; Ares-Mazás, Elvira; Cacciò, Simone M; Gómez-Couso, Hipólito
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.
This is the first report of a blue autofluorescence as a useful characteristic in the microscopic identification of Toxoplasma gondii oocysts. This autofluorescence appears to be of high intensity. Similar to the autofluorescence of related coccidia, the oocysts glow pale blue ...
Ehsan, Amimul; Geurden, Thomas; Casaert, Stijn; Paulussen, Jef; De Coster, Lut; Schoemaker, Toon; Chalmers, Rachel; Grit, Grietje; Vercruysse, Jozef; Claerebout, Edwin
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 (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.
Petersen, Heidi H.; Jianmin, Wang; Mejer, Helena
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......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 environment and is resistant to many common disinfectants. In order to estimate the prevalence of the zoonotic parasite Cryptosporidium in organic pigs and to improve our knowledge of the parasite epidemiology, the prevalence was monitored four times between September 2011 and June 2012 in three Danish...... organic pig farms. Faecal samples were collected for examination of Cryptosporidium spp. with a total of 994 pigs grouped as sows, fatteners, young pigs and piglets. The number of pigs in each age group was 298, 232, 315 and 161 respectively, distributed on the three farms. Faecal samples were collected...
Ghafari, Reza; Rafiei, Abdollah; Tavalla, Mehdi; Moradi Choghakabodi, Parastoo; Nashibi, Rohangez; Rafiei, Reza
This study aimed to determine the prevalence and species of Cryptosporidium among HIV/AIDS patients in southwest of Iran. Two hundred fifty faecal samples from HIV patients were examined for the presence of Cryptosporidium oocysts using a conventional coproscopic approach. Such oocysts were detected in 18 (7.2%) out of 250 faecal samples. Genomic DNAs from 250 samples were then subjected to a nested-PCR-RFLP technique targeting different loci of 18S rRNA gene for species identification. Out of 250 samples, 27 (10.8%) were positive for different Cryptosporidium spp; Restriction patterns resulting from the digestion of the nested amplicon with restriction endonucleases VspI and SspI showed that C. parvum (70.38%) was the most prevalent species, followed by C. hominis (25.92%) and C. meleagridis (3.7%), respectively. The mean CD4+ T-cell count was 215 cells/μL. There was a strong association between cryptosporidiosis and CD4+ T-cell count (P = 0.000) with the highest prevalence recorded among patients with CD4+ T-cell count CD4+ T-cell count increases. Also HIV infection increased the risk of having Cryptosporidium. Our epidemiological findings are useful for any preventive intervention to control disease diffusion. Copyright © 2018 Elsevier Ltd. All rights reserved.
Langkjær, Rikke Breinhold; Vigre, Håkan; Enemark, Heidi L.
The genetic diversity of Cryptosporidium spp. and Giardia duodenalis from dairy cattle and pigs in Denmark was determined in the present study. Faecal samples from 1237 pigs and 1150 cattle originating from 50 sow herds and 50 dairy herds, respectively, were analysed for the presence of the two...... parasites by immunofluorescence microscopy. A large proportion of the (oo)cyst containing samples were selected for molecular characterization. Sequencing and phylogenetic analysis of the 18S rDNA locus and/or the HSP70 gene of 183 pig and 154 cattle isolates of Cryptosporidium revealed the presence of C....... 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...
Silvia Cristina Osaki
Full Text Available Introduction Cryptosporidium is an important protozoan cause of waterborne disease worldwide of concern to public health authorities. To prevent outbreaks of cryptosporidiosis, the monitoring of this parasite in drinking water is necessary. In the present work, the polymerase chain reaction (PCR and nested-PCR techniques were used to detect Cryptosporidium in raw water from catchment points of four water treatment plants (WTP in Curitiba, Paraná, Brazil. Methods First, DNA extraction techniques were tested in samples containing decreasing amount of oocysts in reagent water, and PCR and nested-PCR with specific primers for 18SSU rDNA of Cryptosporidium were conducted to determine their sensitivity. In reagent water, a commercial extraction kit provided the best analytical sensitivity, and PCR and nested-PCR allowed the detection of five and two oocysts, respectively, with the primers XIAOR/XIAOF and XIAO1F/XIAO2R. Results In the spiking experiments, only the PCR with the primers AWA995F/AWA1206R was successful at detecting concentrations of 0.1 oocysts/mL. Two catchments samples of raw water and/or water sludge from four WTPs were contaminated with Cryptosporidium. Conclusions The application of the techniques to monitor Cryptosporidium in water and detect contamination in water catchments of WTPs in Curitiba are discussed in the present work.
Lapen, D R; Schmidt, P J; Thomas, J L; Edge, T A; Flemming, C; Keithlin, J; Neumann, N; Pollari, F; Ruecker, N; Simhon, A; Topp, E; Wilkes, G; Pintar, K D M
Many Cryptosporidium species/genotypes are not considered infectious to humans, and more realistic estimations of seasonal infection risks could be made using human infectious species/genotype information to inform quantitative microbial risk assessments (QMRA). Cryptosporidium oocyst concentration and species/genotype data were collected from three surface water surveillance programs in two river basins [South Nation River, SN (2004-09) and Grand River, GR (2005-13)] in Ontario, Canada to evaluate seasonal infection risks. Main river stems, tributaries, agricultural drainage streams, water treatment plant intakes, and waste water treatment plant effluent impacted sites were sampled. The QMRA employed two sets of exposure data to compute risk: one assuming all observed oocysts were infectious to humans, and the other based on the fraction of oocysts that were C. hominis and/or C. parvum (dominant human infectious forms of the parasite). Viability was not considered and relative infection risk was evaluated using a single hypothetical recreational exposure. Many sample site groupings for both river systems, had significant seasonality in Cryptosporidium occurrence and concentrations (p ≤ 0.05); occurrence and concentrations were generally highest in autumn for SN, and autumn and summer for GR. Mean risk values (probability of infection per exposure) for all sites combined, for each river system, were roughly an order of magnitude lower (avg. of SN and GR 5.3 × 10 -5 ) when considering just C. parvum and C. hominis oocysts, in relation to mean infection risk (per exposure) assuming all oocysts were infectious to humans (5.5 × 10 -4 ). Seasonality in mean risk (targeted human infectious oocysts only) was most strongly evident in SN (e.g., 7.9 × 10 -6 in spring and 8.1 × 10 -5 in summer). Such differences are important if QMRA is used to quantify effects of water safety/quality management practices where inputs from a vast array of fecal pollution
Hohweyer, Jeanne; Cazeaux, Catherine; Travaillé, Emmanuelle; Languet, Emilie; Dumètre, Aurélien; Aubert, Dominique; Terryn, Christine; Dubey, Jitender P; Azas, Nadine; Houssin, Maryline; Loïc, Favennec; Villena, Isabelle; La Carbona, Stéphanie
Toxoplasma gondii, Cryptosporidium spp. and Giardia intestinalis are emerging pathogen parasites in the food domain. However, without standardized methods for their detection in food matrices, parasitic foodborne outbreaks remain neglected. In this study, a new immunomagnetic separation assay (IMS Toxo) targeting the oocyst's wall of T. gondii was developed using a specific purified monoclonal antibody. Performance of this IMS Toxo coupled to microscopic and qPCR analyses was evaluated in terms of limit of detection (LOD) and recovery rate (RR) on: i) simple matrix (LOD = 5 oocysts; RR between 5 and 56%); ii) raspberries and basil (LOD = 33 oocysts/g; RR between 0.2 and 35%). Finally, to simultaneously extract the three protozoa from these food matrices, T. gondii oocysts were directly concentrated (without IMS Toxo) from the supernatant of the IMS of Cryptosporidium and Giardia (oo)cysts. This strategy associated to qPCR detection led to LOD <1 to 3 (oo)cysts/g and RR between 2 and 35%. This procedure was coupled to RT-qPCR analyses and showed that the three protozoa persisted on the leaves of basil and remained viable following storage at 4 °C for 8 days. These data strengthen the need to consider these protozoa in food safety. Copyright © 2016 Elsevier Ltd. All rights reserved.
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
Margareth Leitão Gennari-Cardoso
Full Text Available This study's objective was to search for Cryptosporidium sp. in diarrheic feces from children aged zero to 12 years and cared for at medical units within Universidade Federal de Uberlândia or at a private practice in Uberlândia, State of Minas Gerais, Brazil, from September 1992 to August 1993. Three fecal samples preserved in 10% formalin, were collected from 94 children. Oocyst concentration was performed through Ritchie's (modified method and staining of fecal smears for each sample (total of 1128 slides was done by the "Safranin/Methylene Blue" and the "Kinyoun (modified" techniques. The Hoffmann, Pons & Janer method was also employed to look for other enteroparasites. From 94 children, 4.26% excreted fecal Cryptosporidium oocysts. The infection seemed to vary according to age: 5.08% of patients aged zero to two years old; 33.33% of those aging eight to ten years (P>0.05. Cryptosporidium appeared in November, December and March, during the rainy season. 20.21% of the children harbored at least one enteroparasite different from Cryptosporidium, mainly Giardia intestinalis (12.77%. From Cryptosporidium infected patients, two had only this kind, another harbored Giardia intestinalis; the last one hosted Strongyloides stercoralis.
Ifeonu, Olukemi O.; Chibucos, Marcus C.; Orvis, Joshua; Su, Qi; Elwin, Kristin; Guo, Fengguang; Zhang, Haili; Xiao, Lihua; Sun, Mingfei; Chalmers, Rachel M.; Fraser, Claire M.; Zhu, Guan; Kissinger, Jessica C.; Widmer, Giovanni; Silva, Joana C.
Human cryptosporidiosis is caused primarily by Cryptosporidium hominis, C. parvum and C. meleagridis. To accelerate research on parasites in the genus Cryptosporidium, we generated annotated, draft genome sequences of human C. hominis isolates TU502_2012 and UKH1, C. meleagridis UKMEL1, also isolated from a human patient, and the avian parasite C. baileyi TAMU-09Q1. The annotation of the genome sequences relied in part on RNAseq data generated from the oocyst stage of both C. hominis and C. b...
Zhang, Weizhe; Shen, Yujuan; Wang, Rongjun; Liu, Aiqin; Ling, Hong; Li, Yihong; Cao, Jianping; Zhang, Xiaoyun; Shu, Jing; Zhang, Longxian
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. 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. 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.
.6%. Sensitivity and specificity of coproantigen detection compared with the microscopic was 100% and 71.4%. Detection coproantigen necessary in patients with high suspicion of cryptosporidiosis but oocysts detection was negative. Key words: Cryptosporidium sp, HIV/AIDS, chronic diarrhea, coproantigen
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.
Matheus Hillard Farret
Full Text Available This study reports the gastrointestinal parasitism by Giardia sp. and Cryptosporidium sp. in blue macaw (Anodorhynchus hyacinthinus in the southern region of Brazil. Fecal samples of two species kept in captivity in the state of Rio Grande do Sul were analyzed by the direct smear method, centrifugal flotation technique with zinc sulfate and Kinyoun staining technique for research of parasites. Mixed infection by eggs of Capillaria, cysts of Giardia sp. and oocysts of Cryptosporidium sp. was observed. This is the first report this protozoa in blue macaw.
García-Presedo, Ignacio; Pedraza-Díaz, Susana; González-Warleta, Marta; Mezo, Mercedes; Gómez-Bautista, Mercedes; Ortega-Mora, Luis Miguel; Castro-Hermida, José Antonio
The aim of the present study was to identify the species and/or genotypes of Cryptosporidium and Giardia duodenalis infecting roe deer (Capreolus capreolus) in Galicia (NW Spain). The presence of both enteropathogens was investigated in 212 faecal samples from roe deer shot in diverse game preserves in three different areas of Galicia. The samples were analyzed by immunofluorescence microscopy and PCR amplification, and fragments of the 18S SSU rRNA gene of Cryptosporidium and the β-giardin gene of G. duodenalis were sequenced. In total, 9 samples (4.2%) were positive for Cryptosporidium and 19 samples (8.9%) for G. duodenalis. These samples tested positive with both techniques. However, gene sequencing was only possible for Cryptosporidium in 6 of the samples and for G. duodenalis in 7 of the samples. Cryptosporidium bovis was identified in 3 samples and C. ryanae oocysts were detected in another 3 samples. Sequencing of the amplicons identified G. duodenalis sub-assemblage A-II in 7 samples. Both Cryptosporidium and G. duodenalis infections were more prevalent in juvenile than in adult animals, although the differences were not significant. G. duodenalis was more prevalent than Cryptosporidium in both age groups, although again the differences were not statistically significant. The mean intensity of infection by Cryptosporidium and G. duodenalis was similar in both age groups and ranged between 5 and 225 oocysts/g and 5 and 320 cysts/g of faeces, respectively. This study represents the first molecular characterization of these parasites in Spanish roe deer. Identification of C. bovis and G. duodenalis sub-assemblage A-II indicates that zoonotic transmission of these enteropathogens between roe deer and humans is possible and that cross transmission of some Cryptosporidium species and G. duodenalis (sub-assemblage A-II) may occur between related animal species sharing the same habitats. Copyright © 2013 Elsevier B.V. All rights reserved.
Zahedi, Alireza; Monis, Paul; Aucote, Sarah; King, Brendon; Paparini, Andrea; Jian, Fuchun; Yang, Rongchang; Oskam, Charlotte; Ball, Andrew; Robertson, Ian; Ryan, Una
Cryptosporidium is one of the most common zoonotic waterborne parasitic diseases worldwide and represents a major public health concern of water utilities in developed nations. As animals in catchments can shed human-infectious Cryptosporidium oocysts, determining the potential role of animals in dissemination of zoonotic Cryptosporidium to drinking water sources is crucial. In the present study, a total of 952 animal faecal samples from four dominant species (kangaroos, rabbits, cattle and sheep) inhabiting Sydney's drinking water catchments were screened for the presence of Cryptosporidium using a quantitative PCR (qPCR) and positives sequenced at multiple loci. Cryptosporidium species were detected in 3.6% (21/576) of kangaroos, 7.0% (10/142) of cattle, 2.3% (3/128) of sheep and 13.2% (14/106) of rabbit samples screened. Sequence analysis of a region of the 18S rRNA locus identified C. macropodum and C. hominis in 4 and 17 isolates from kangaroos respectively, C. hominis and C. parvum in 6 and 4 isolates respectively each from cattle, C. ubiquitum in 3 isolates from sheep and C. cuniculus in 14 isolates from rabbits. All the Cryptosporidium species identified were zoonotic species with the exception of C. macropodum. Subtyping using the 5' half of gp60 identified C. hominis IbA10G2 (n = 12) and IdA15G1 (n = 2) in kangaroo faecal samples; C. hominis IbA10G2 (n = 4) and C. parvum IIaA18G3R1 (n = 4) in cattle faecal samples, C. ubiquitum subtype XIIa (n = 1) in sheep and C. cuniculus VbA23 (n = 9) in rabbits. Additional analysis of a subset of samples using primers targeting conserved regions of the MIC1 gene and the 3' end of gp60 suggests that the C. hominis detected in these animals represent substantial variants that failed to amplify as expected. The significance of this finding requires further investigation but might be reflective of the ability of this C. hominis variant to infect animals. The finding of zoonotic Cryptosporidium species in these animals may
Full Text Available Cryptosporidium is one of the most common zoonotic waterborne parasitic diseases worldwide and represents a major public health concern of water utilities in developed nations. As animals in catchments can shed human-infectious Cryptosporidium oocysts, determining the potential role of animals in dissemination of zoonotic Cryptosporidium to drinking water sources is crucial. In the present study, a total of 952 animal faecal samples from four dominant species (kangaroos, rabbits, cattle and sheep inhabiting Sydney's drinking water catchments were screened for the presence of Cryptosporidium using a quantitative PCR (qPCR and positives sequenced at multiple loci. Cryptosporidium species were detected in 3.6% (21/576 of kangaroos, 7.0% (10/142 of cattle, 2.3% (3/128 of sheep and 13.2% (14/106 of rabbit samples screened. Sequence analysis of a region of the 18S rRNA locus identified C. macropodum and C. hominis in 4 and 17 isolates from kangaroos respectively, C. hominis and C. parvum in 6 and 4 isolates respectively each from cattle, C. ubiquitum in 3 isolates from sheep and C. cuniculus in 14 isolates from rabbits. All the Cryptosporidium species identified were zoonotic species with the exception of C. macropodum. Subtyping using the 5' half of gp60 identified C. hominis IbA10G2 (n = 12 and IdA15G1 (n = 2 in kangaroo faecal samples; C. hominis IbA10G2 (n = 4 and C. parvum IIaA18G3R1 (n = 4 in cattle faecal samples, C. ubiquitum subtype XIIa (n = 1 in sheep and C. cuniculus VbA23 (n = 9 in rabbits. Additional analysis of a subset of samples using primers targeting conserved regions of the MIC1 gene and the 3' end of gp60 suggests that the C. hominis detected in these animals represent substantial variants that failed to amplify as expected. The significance of this finding requires further investigation but might be reflective of the ability of this C. hominis variant to infect animals. The finding of zoonotic Cryptosporidium species in these
Lee, Sangun; Harwood, Melanie; Girouard, Don; Meyers, Marvin J; Campbell, Mary A; Beamer, Gillian; Tzipori, Saul
Recent reports highlighting the global significance of cryptosporidiosis among children, have renewed efforts to develop control measures. We have optimized the gnotobiotic piglet model of acute diarrhea to evaluate azithromycin (AZR), nitazoxanide (NTZ), or treatment with both against Cryptosporidium hominis, the species responsible for most human cases. Piglets, animals reproducibly clinically susceptible to C. hominis, when inoculated with 106 oocysts, developed acute diarrhea with oocyst excretion in feces within 3 days. Ten day-treatment with recommended doses for children, commencing at onset of diarrhea, showed that treatment with AZR or NTZ relieved symptoms early in the treatment compared with untreated animals. Piglets treated with AZR exhibited no reduction of oocyst excretion whereas treatment with NTZ significantly reduced oocyst shedding early, increasing however after 5 days. While treatment with AZR+NTZ led to considerable symptomatic improvement, it had a modest effect on reducing mucosal injury, and did not completely eliminate oocyst excretion. Doubling the dose of AZR and/or NTZ did not improve the clinical outcome, confirming clinical observations that NTZ is only partially effective in reducing duration of diarrhea in children. This investigation confirms the gnotobiotic piglet as a useful tool for drug evaluation for the treatment of cryptosporidiosis in children.
Koudela, Břetislav; Modrý, D.
Roč. 45, č. 1 (1998), s. 8 ISSN 1066-5234. [ Cryptosporidium sp. in lazards. 01.01.1998-02.01.1998, Praha] R&D Projects: GA ČR GA508/95/0273; GA AV ČR IPP2020702 Subject RIV: fp - Other Medical Disciplines
Genotypic identification of Cryptosporidium spp. isolated from hiv-infected patients and immunocompetent children of São Paulo, Brazil Identificação genotípica de Cryptosporidium spp. isolados a partir de pacientes com HIV e crianças imunocompetentes de São Paulo, Brasil
Ana Julia Urias dos Santos Araújo
Full Text Available Cryptosporidium isolates identified in fourteen stool samples, collected from five HIV-infected patients and nine immunocompetent children, living in the Sate of São Paulo, Brazil, were submitted to a molecular analysis using a nested PCR followed of restriction fragment length polymorphism (RFLP, for genetic characterization. The analysis was based on digestion with RsaI restriction enzyme of a DNA fragment amplified from the Cryptosporidium oocyst wall protein (COWP gene. Based on this analysis, four samples were identified as Cryptosporidium parvum, eight as Cryptosporidium hominis and two presented a profile that correspondedto Cryptosporidium meleagridis when compared to the standards used in the analysis. The use of molecular methods can be helpful to identify source of infections and risk factors related to Cryptosporidium infection in our communities.Isolados de Cryptosporidium identificados em quatorze amostras de fezes, coletadas de cinco pacientes com infecção por HIV e de nove crianças imunocompetentes, residentes no estado de São Paulo, Brasil, foram submetidos a análise molecular por Nested-PCR, seguido da caracterização genética por polimorfismo do tamanho do fragmento de restrição (RFLP. A análise foi baseada na digestão, com a enzima de restrição RsaI, de um fragmento de DNA amplificado do gene que codifica a proteína de parede do oocisto de Cryptosporidium (COWP. Baseado nesta análise, quando comparadas aos padrões utilizados, quatro amostras foram identificadas como Cryptosporidium parvum, oito como Cryptosporidium hominis e duas apresentaram um perfil correspondente ao de Cryptosporidium meleagridis. O uso de métodos moleculares pode ser útil para identificar a fonte das infecções e os fatores de risco relacionados à infecção por Cryptosporidium em nossas comunidades.
Robin, G; Fraser, D; Orr, N; Sela, T; Slepon, R; Ambar, R; Dagan, R; Le Blancq, S; Deckelbaum, R J; Cohen, D
An enzyme-linked immunosorbent assay system using oocyst lysate as antigen was used to detect serum- specific antibody responses to Cryptosporidium parvum between 1989 and 1994 in consecutive sera obtained at birth, and at the age of 6, 12, and 23 months, from 52 infants living in a Bedouin town located in the south of Israel. The serologic tests revealed high levels of immunoglobulin G anti-Cryptosporidium at birth that dropped significantly by the age of 6 months and then rose continuously to a geometric mean titer of 481 at age 23 months. The serum immunoglobulin M Cryptosporidium antibodies rose continuously from nearly undetectable levels at birth to a geometric mean titer of 471 (157-fold increase) at age 23 months. All the subjects already showed at 6 months a significant rise in immunoglobulin M. A significant rise in immunoglobulin A titers was detected in 48% and 91% of subjects at 6 and 23 months, respectively. By monthly surveillance, microscopy using the modified Ziehl-Neelsen method and confirmed by indirect immunofluorescence assay detected Cryptosporidium antigens in only 11% at age 6 months and 48% at age 23 months. The extent of exposure to Cryptosporidium immediately after birth as detected by serology is much higher than that predicted by frequent prospective assessment of stool samples.
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
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.
Appelbee, A J; Thompson, R C A; Measures, L M; Olson, M E
Giardia and Cryptosporidium are protozoan parasites known to cause enteric disease in terrestrial wildlife species (mammals, reptiles and birds). Few surveys for Giardia and Cryptosporidium in marine wildlife species, such as pinnipeds, have been reported. The objective of this study was to determine the prevalence and genotype of Giardia and Cryptosporidium in two species of pinnipeds, harp seal (Phoca groenlandica) and hooded seal (Cystophora cristata), from the Gulf of St. Lawrence, Canada. Faecal samples were collected from pup and adult seals and examined for the presence of cysts of Giardia and oocysts of Cryptosporidium using microscopy and immunofluorescent staining. Tissues from the small intestine of adult seals were also collected and examined for infections using the polymerase chain reaction (PCR) technique. Giardia cysts were found in the faeces of 42% (16/38) of adult harp seals, but in none of the harp seal pups (0/20). Although Giardia cysts were not detected in faeces of adult hooded seals (0/10) using microscopy, 80% tested positive for Giardia using PCR of intestinal tissue indicative of a true replicating infection. Both harp and hooded seals harboured infections with the zoonotic strain, Giardia duodenalis Assemblage A, as determined using a nested-PCR technique to amplify a small subunit ribosomal (SSU-rRNA) gene of Giardia. Cryptosporidium was not detected by microscopy, nor using the PCR technique on intestinal tissues from any of the 68 seals examined. Copyright 2010 Elsevier B.V. All rights reserved.
Moreira, Maria João; Soares, Sónia; de Lurdes Delgado, Maria; Figueiredo, João; Silva, Elisabete; Castro, António; Cosa, José Manuel Correida Da
Cryptosporidium and Giardia are 2 protozoan parasites responsible for waterborne diseases outbreaks worldwide. In order to assess the prevalence of these protozoans in drinking water samples in the northern part of Portugal and the risk of human infection, we have established a long term program aiming at pinpointing the sources of surface water, drinking water, and environmental contamination, working with the water-supply industry. Total 43 sources of drinking water samples were selected, and a total of 167 samples were analyzed using the Method 1623. Sensitivity assays regarding the genetic characterization by PCR and sequencing of the genes, 18S SSU rRNA, for Cryptosporidium spp. and β,-giardin for G. duodenalis were set in the laboratory. According to the defined criteria, molecular analysis was performed over 4 samples. Environmental stages of the protozoa were detected in 25.7% (43 out of 167) of the water samples, 8.4% (14 out of 167) with cysts of Giardia, 10.2% (17 out of 167) with oocysts of Cryptosporidium and 7.2% (12 out of 167) for both species. The mean concentrations were 0.1-12.7 oocysts of Cryptosporidium spp. per 10 L and 0.1-108.3 cysts of Giardia duodenalis per 10 L. Our results suggest that the efficiency in drinking water plants must be ameliorated in their efficiency in reducing the levels of contamination. We suggest the implementation of systematic monitoring programs for both protozoa. To authors' knowledge, this is the first report evaluating the concentration of environmental stages of Cryptosporidium and Giardia in drinking water samples in the northern part of Portugal. PMID:20333284
Full Text Available Cryptosporidiosis has recently attracted attention as an emerging waterborne and foodborne disease as well as an opportunistic infection in HIV infected individuals. The lack of genetic information, however, has resulted in confusion in the taxonomy of Cryptosporidium parasites and in the development of molecular tools for the identification and typing of oocysts in environmental samples. Phylogenetic analysis of the small subunit ribosomal RNA (SSU rRNA gene has shown that the genus Cryptosporidium is comprised of several distinct species. Our data show the presence of at least four species: C. parvum, C. muris, C. baileyi and C. serpentis (C. meleagridis, C. nasorum and C. felis were not studied. Within each species, there is some sequence variation. Thus, various genotypes (genotype 1, genotype 2, guinea pig genotype, monkey genotype and koala genotype, etc. of C. parvum differ from each other in six regions of the SSU rRNA gene. Information on polymorphism in Cryptosporidium parasites has been used in the development of species and strain-specific diagnostic tools. Use of these tools in the characterization of oocysts various samples indicates that C. parvum genotype 1 is the strain responsible for most human Cryptosporidium infections. In contrast, genotype 2 is probably the major source for environmental contamination of environment, and has been found in most oysters examined from Chesapeake Bay that serve as biologic monitors of surface water. Parasites of Cryptosporidium species other than C. parvum have not been detected in HIV+ individuals, indicating that the disease in humans is caused only by C. parvum.
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.
Almeida, André; Moreira, Maria João; Soares, Sónia; Delgado, Maria de Lurdes; Figueiredo, João; Silva, Elisabete; Castro, António; Cosa, José Manuel Correida Da
Cryptosporidium and Giardia are 2 protozoan parasites responsible for waterborne diseases outbreaks worldwide. In order to assess the prevalence of these protozoans in drinking water samples in the northern part of Portugal and the risk of human infection, we have established a long term program aiming at pinpointing the sources of surface water, drinking water, and environmental contamination, working with the water-supply industry. Total 43 sources of drinking water samples were selected, and a total of 167 samples were analyzed using the Method 1623. Sensitivity assays regarding the genetic characterization by PCR and sequencing of the genes, 18S SSU rRNA, for Cryptosporidium spp. and beta,-giardin for G. duodenalis were set in the laboratory. According to the defined criteria, molecular analysis was performed over 4 samples. Environmental stages of the protozoa were detected in 25.7% (43 out of 167) of the water samples, 8.4% (14 out of 167) with cysts of Giardia, 10.2% (17 out of 167) with oocysts of Cryptosporidium and 7.2% (12 out of 167) for both species. The mean concentrations were 0.1-12.7 oocysts of Cryptosporidium spp. per 10 L and 0.1-108.3 cysts of Giardia duodenalis per 10 L. Our results suggest that the efficiency in drinking water plants must be ameliorated in their efficiency in reducing the levels of contamination. We suggest the implementation of systematic monitoring programs for both protozoa. To authors' knowledge, this is the first report evaluating the concentration of environmental stages of Cryptosporidium and Giardia in drinking water samples in the northern part of Portugal.
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
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
Stephanie J Salyer
Full Text Available Cryptosporidium is one of the most common parasitic diarrheal agents in the world and is a known zoonosis. We studied Cryptosporidium in people, livestock, and non-human primates in the region of Kibale National Park, Uganda. Land use change near the park has resulted in fragmented forest patches containing small, remnant populations of wild primates that interact intensively with local people and livestock. Our goal was to investigate risk factors for Cryptosporidium infection and to assess cross-species transmission using molecular methods.Diagnostic PCR revealed a prevalence of Cryptosporidium of 32.4% in humans, 11.1% in non-human primates, and 2.2% in livestock. In the case of humans, residence in one particular community was associated with increased risk of infection, as was fetching water from an open water source. Although 48.5% of infected people reported gastrointestinal symptoms, this frequency was not significantly different in people who tested negative (44.7% for Cryptosporidium, nor was co-infection with Giardia duodenalis associated with increased reporting of gastrointestinal symptoms. Fecal consistency was no different in infected versus uninfected people or animals. DNA sequences of the Cryptosporidium oocyst wall protein gene placed all infections within a well-supported C. parvum/C. hominis clade. However, the only two sequences recovered from primates in the core of the park's protected area fell into a divergent sub-clade and were identical to published sequences from C. parvum, C. hominis, and C. cuniculus, suggesting the possibility of a separate sylvatic transmission cycle.Cryptosporidium may be transmitted frequently among species in western Uganda where people, livestock, and wildlife interact intensively as a result of anthropogenic changes to forests, but the parasite may undergo more host-specific transmission where such interactions do not occur. The parasite does not appear to have strong effects on human or
Vermeulen, Lucie C.; de Kraker, Jelske; Hofstra, Nynke; Kroeze, Carolien; Medema, Gertjan
Cryptosporidium is a protozoan parasite that can cause diarrhoea. Human faeces are an important source of Cryptosporidium in surface waters. We present a model to study the impact of sanitation, urbanization and population growth on human emissions of Cryptosporidium to surface waters. We build on a global model by Hofstra et al (2013 Sci. Total Environ. 442 10-9) and zoom into Bangladesh and India as illustrative case studies. The model is most sensitive to changes in oocyst excretion and infection rate, and to assumptions on the share of faeces reaching the surface water for different sanitation types. We find urban centres to be hotspots of human Cryptosporidium emissions. We estimate that 53% (Bangladesh) and 91% (India) of total emissions come from urban areas. 50% of oocysts come from only 8% (Bangladesh) and 3% (India) of the country area. In the future, population growth and urbanization may further deteriorate water quality in Bangladesh and India, despite improved sanitation. Under our ‘business as usual’ (‘sanitation improvements’) scenario, oocyst emissions will increase by a factor 2.0 (1.2) for India and 2.9 (1.1) for Bangladesh between 2010 and 2050. Population growth, urbanization and sanitation development are important processes to consider for large scale water quality modelling.
Feng, Yaoyu; Li, Na; Duan, Liping; Xiao, Lihua
To identify the genotype and subtype distributions of Cryptosporidium oocysts in domestic wastewater in Shanghai, China, and to facilitate the characterization of the endemic transmission of cryptosporidiosis, raw domestic wastewater samples were collected from four wastewater treatment plants in Shanghai, China, from December 2006 to April 2007. Genotypes of Cryptosporidium species were detected based on PCR-restriction fragment length polymorphism and sequence analyses of the small-subunit rRNA gene. Samples that contained Cryptosporidium hominis were further subtyped by DNA sequencing of the 60-kDa glycoprotein gene. Among a total of 90 samples analyzed, 63 were PCR positive, 10 of which had mixed genotypes. Fifty-nine (93.7%) of the PCR-positive samples had C. hominis, and 7 (11.1%) had C. meleagridis. The other seven Cryptosporidium species/genotypes identified included C. baileyi, C. parvum, C. suis, C. muris, rat genotype, avian genotype III, and a novel genotype. Forty-seven of the 59 C. hominis-positive samples were successfully subtyped, with 29 having subtype family Ib and the remaining belonging to subtype families Ia, Id, Ie, and If. The three Ib subtypes identified, IbA19G2, IbA20G2, and IbA21G2, were very different from the two common Ib subtypes (IbA9G3 and IbA10G2) found in other areas of the world. Likewise, the Ie subtype IeA12G3T3 was also different from the common IeA11G3T3 subtype. Thus, the presence of multiple subtype families and unique Ib, Ie, and If subtypes indicates that there might be endemic transmission of cryptosporidiosis in the study area and that C. hominis populations there might be very different from those in other areas.
Gómez-Couso, H; Fontán-Sainz, M; Fernández-Alonso, J; Ares-Mazás, E
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.
Full Text Available Aim: Cryptosporidiosis is a worldwide zoonosis. Microscopic examinations may fail due to indistinctive morphological peculiarities of causative species. Hence, molecular diagnostics has become more important. Methods: Stool samples from 150 patients were examined using carbol-fuchsin stain to determine Cryptosporidium spp. oocysts. Combined nested polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP technique was used for establishing different species in positive samples. The samples were also screened for other parasites by wet-mount and zinc sulfate flotation methods. Results: Microscopic examinations and molecular techniques revealed 0.67% (1/150 and 8.93% (5/56 positivity, respectively. Nested PCR-RFLP enabled the detection of Cryptosporidium hominis (C. hominis in one sample, while Cryptosporidium parvum (C. parvum was detected in four samples. With this study, C. hominis was reported from humans for the first time in Turkey. Among infected ones, three of which were children, four patients excreted C. parvum oocysts had gastroenteritis, and a patient positive for C. hominis had gastroenteritis accompanied by nausea and vomiting. No Giardia spp. and Entamoeba spp. were detected in all infected individuals. Conclusion: C. parvum cases outnumbered C. hominis cases, suggesting a zoonotic transmission although infected individuals were living in an urban area where animal husbandry was not allowed. However, water-borne pathogen contamination in the city’s water supply is considered a factor for transmission.
Asadpour, Mohammad; Namazi, Fatemeh; Razavi, Seyed Mostafa; Nazifi, Saeed
Cryptosporidium is a ubiquitous protozoan parasite causing gastrointestinal disorder in various hosts worldwide. The disease is self-limiting in the immunocompetent but life-threatening in immunodeficient individuals. Investigations to find an effective drug for the complete elimination of the Cryptosporidium infection are ongoing and urgently needed. The current study was undertaken to examine the anti-cryptosporidial efficacy of curcumin in experimentally infected mice compared with that of paromomycin. Oocysts were isolated from a pre-weaned dairy calf and identified as Cryptosporidium parvum using a nested- polymerase chain reaction (PCR) on Small subunit ribosomal ribonucleic acid (SSU rRNA) gene and sequencing analysis. One hundred and ten female BALB/c mice were divided into five groups. Group 1 was infected and treated with curcumin; Group 2 infected and treated with paromomycin; Group 3 infected without treatment; Group 4 included uninfected mice treated with curcumin, and Group 5 included uninfected mice treated with distilled water for 11 successive days, starting on the first day of oocyst shedding. The oocyst shedding was recorded daily. At days 0, 3, 7, and 11 of post treatments, five mice from each group were killed humanly; jejunum and ileum tissue samples were processed for histopathological evaluation and counting of oocyst on villi, simultaneously. Furthermore, total antioxidant capacity (TAC) and malondialdehyde (MDA) concentrations in affected tissues were also measured in different groups. By treatments, tissue lesions and the number of oocyst on villi of both jejunum and ileum were decreased with a time-dependent manner. In comparison with Group 3, oocyst shedding was stopped at the end of treatment period in both groups 1 and 2 without recurrence at 10days after drug withdrawal. Also, TAC was increased and the MDA concentrations were decreased in Group 1. Moreover, paromomycin showed acceptable treatment outcomes during experiment and its
Magana-Ordorica, Dalia; Mena, Kristina; Valdez-Torres, Jose B; Soto-Beltran, Marcela; Leon-Felix, Josefina; Chaidez, Cristobal
Untreated sewage has adversely affected the quality of marine recreational waters worldwide. Exposure to marine recreational water with poor microbial quality may pose a threat to bathers. The objectives of this study were to assess the effect of physicochemical parameters on Cryptosporidium and Giardia presence in marine recreational water of Sinaloa, Mexico, by Logistic Regression Analyses. Thirty-two 10-litre water samples were collected from two tourist beaches, Altata and Mazatlan, between November 2006 and May 2007. Water samples were processed by the EPA 1623 method and pH, temperature, salinity and turbidity were also determined. Cryptosporidium and Giardia were present in 71 and 57% of the samples collected from Altata, respectively. In Mazatlan, Cryptosporidium and Giardia were found in 83 and 72% of the samples, respectively. The overall concentration of Cryptosporidium ranged from 150 to 2,050 oocysts/10 L with an average of 581 oocysts/10 L and Giardia ranged from 10 to 300 cysts/10 L with an average of 73 cysts/10 L. The occurrence of both parasites increased in water with decreasing temperatures and increasing turbidity of the water.
Emanuel S. Swai
Full Text Available A cross-sectional study was conducted to determine prevalence and risk factors of cryptosporidiosis in bovine from two contrasting production system in and around Tanga municipality between May 2003 and January 2004. The study populations comprised 117 calves aged ≤3 months, randomly selected from 44 smallholders dairy and traditional managed herds, respectively. Individual calf and herd-level information was collected using a structured questionnaire and feacal samples were screened for Cryptosporidium spp oocysts using the modified Ziehl-Neelsen method. Overall, 35% of the calves in the study were shedding Cryptosporidium spp oocysts, with at least one positive calf detected in 54.5% of herds. Independent risk factors for cryptosporidiosis were: age ≥1 to ≤2 months and level of cleanness of calf house floor categorized as dirty (<.05. Similarly an increases risk of Cryptosporidium spp infection was found in calves from smallholder dairy units compared to traditional herds (<.05. The finding highlights that Cryptosporidium spp is prevalent among calves in the area under study. The high prevalence of cryptosporidiosis detected in this study suggests that it may have a significant impact on livestock industry and that the close interaction between cattle and human may play a role in zoonotic transmission to humans.
Farkas, K; Plutzer, J; Moltchanova, E; Török, A; Varró, M J; Domokos, K; Frost, F; Hunter, P R
In this study the putative protective seroprevalence (PPS) of IgG antibodies to the 27-kDa and 15/17-kDa Cryptosporidium antigens in sera of healthy participants who were and were not exposed to Cryptosporidium oocysts via surface water-derived drinking water was compared. The participants completed a questionnaire regarding risk factors that have been shown to be associated with infection. The PPS was significantly greater (49-61%) in settlements where the drinking water originated from surface water, than in the control city where riverbank filtration was used (21% and 23%). Logistic regression analysis on the risk factors showed an association between bathing/swimming in outdoor pools and antibody responses to the 15/17-kDa antigen complex. Hence the elevated responses were most likely due to the use of contaminated water. Results indicate that waterborne Cryptosporidium infections occur more frequently than reported but may derive from multiple sources.
Deshpande, A; Alexander, C L; Coyne, M; Brownlie, S; Smith-Palmer, A; Jones, B L
Cryptosporidium hominis is one of the most prevalent protozoan parasites to infect humans where transmission is via the consumption of infective oocysts. This study describes sporadic cases in addition to the molecular diversity of outbreak cases in Scotland using the glycoprotein-60 subtyping tool. From a total of 187 C. hominis isolates, 65 were subjected to further molecular analysis and 46 were found to be the common IbA10G2 subtype. Unusual subtypes included four isolates belonging to the Ia family (IaA14R3, n = 12; IaA14R2, n = 1; IaA9G3, n = 1; IaA25R3, n = 2), two from the Id family (IdA24, n = 1; IdA17, n = 1) and one belonging to the Ie family, namely IeA11G3T3. These data contribute significantly to our knowledge and understanding of the molecular diversity of C. hominis isolates from outbreak investigations involving Scottish residents which will be beneficial for the management of future outbreaks.
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
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.
Roberta Flávia Ribeiro Rolando
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.
Toxaplasma gondii is a parasitic protozoan that infects a wide range of vertebrates, including humans. This report describes methods that have been developed for separation of oocyst components starting with the mechanical fragmentation of oocysts. Use of iodixoanol gradients a...
Agnes Kurniawan; Sri W. Dwintasari; Herbowo A. Soetomenggolo; Septelia I. Wanandi
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 wi...
Paiva, Philipp Ricardo S O; Grego, Kathleen F; Lima, Valéria M F; Nakamura, Alex A; da Silva, Deuvânia C; Meireles, Marcelo V
Infection by Cryptosporidium serpentis is one of the most important diseases in reptiles and is characterized by chronic clinical or subclinical infection and the presence of hypertrophic gastritis, food regurgitation, progressive weight loss, mortality, and intermittent or continuous shedding of oocysts in the feces. The objectives of this study were to standardize an indirect enzyme-linked immunosorbent assay (ELISA) to detect antibodies against C. serpentis and to evaluate the clinical, parasitological, and humoral immune response in snakes naturally infected with C. serpentis. Twenty-one snakes naturally infected with C. serpentis and housed at the Butantan Institute, São Paulo, Brazil, underwent clinical and parasitological analyses for C. serpentis infection through daily records of clinical signs and a monthly survey of fecal shedding of oocysts using the Kinyoun's acid-fast staining. The serological evaluation was performed monthly by indirect ELISA using crude total antigen from oocysts of C. serpentis to detect anti-C. serpentis antibodies. Clinical symptoms consisted of food regurgitation, inappetence, and progressive weight loss. The parasitological analysis revealed intermittent fecal shedding of a variable number of oocysts in all snakes, with positivity in 85.32% (157/184) of the samples. The indirect ELISA was positive in 68.25% (86/126) of the samples. A humoral immune response was observed in most animals; however, fluctuating antibodies levels, leading to alternating positive and negative results, were observed in most snakes. Copyright © 2013 Elsevier B.V. All rights reserved.
Borges, João Carlos; Lima, Danielle Dos; da Silva, Edson Moura; Moreira, André Lucas de Oliveira; Marmontel, Miriam; Carvalho, Vitor Luz; Amaral, Rodrigo de; Lazzarini, Stella Maris; Alves, Leucio Câmara
Cryptosporidium and Giardia are protozoans that can infect humans and wild and domestic animals. Due to the growing importance of diseases caused by protozoan parasites in aquatic species, we aimed to evaluate the frequency of infection by Cryptosporidium spp. and Giardia sp. in aquatic and marine mammals in the northern and northeastern regions of Brazil. We collected 553 fecal samples from 15 species of wild-ranging and captive aquatic mammals in northern and northeastern Brazil. All samples were analyzed by the Kinyoun technique for identification of Cryptosporidium spp. oocysts. Giardia sp. cysts were identified by means of the centrifugal-flotation technique in zinc sulfate solution. Subsequently, all samples were submitted for direct immunofluorescence testing. The overall frequency of infection was 15.55% (86/553) for Cryptosporidium spp. and 9.04% (50/553) for Giardia sp. The presence of Cryptosporidium spp. was detected in samples from 5 species: neotropical river otter Lontra longicaudis (15.28%), giant otter Pteronura brasiliensis (41.66%), Guiana dolphin Sotalia guianensis (9.67%), Amazonian manatee Trichechus inunguis (16.03%), and Antillean manatee T. manatus (13.79%). Giardia sp. was identified in L. longicaudis (9.23%), P. brasiliensis (29.16%), pygmy sperm whale Kogia breviceps (100%), dwarf sperm whale K. sima (25%), S. guianensis (9.67%), T. inunguis (3.81%), and T. manatus (10.34%). This is the first report of Cryptosporidium spp. in L. longicaudis, P. brasiliensis, and S. guianensis, while the occurrence of Giardia sp., in addition to the 2 otter species, was also identified in manatees, thus extending the number of hosts susceptible to these parasitic agents.
The ability of microorganisms to survive under extreme conditions is closely related to the physicochemical properties of their wall. In the ubiquitous protozoan parasite Toxoplasma gondii, the oocyst stage possesses a bilayered wall that protects the dormant but potentially infective parasites from...
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
Elenice Messias do Nascimento Gonçalves
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
Full Text Available BACKGROUND: The trehalose synthetic pathway is present in bacteria, fungi, plants and invertebrate animals, but is absent in vertebrates. This disaccharide mainly functions as a stress protectant against desiccation, heat, cold and oxidation. Genes involved in trehalose synthesis have been observed in apicomplexan parasites, but little was known about these enzymes. Study on trehalose synthesis in apicomplexans would not only shed new light into the evolution of this pathway, but also provide data for exploring this pathway as novel drug target. METHODOLOGY/PRINCIPAL FINDINGS: We have observed the presence of the trehalose synthetic pathway in Cryptosporidium and other apicomplexans and alveolates. Two key enzymes (trehalose 6-phosphate synthase [T6PS; EC 18.104.22.168] and trehalose phosphatase [TPase; EC 22.214.171.124] are present as Class II bifunctional proteins (T6PS-TPase in the majority of apicomplexans with the exception of Plasmodium species. The enzyme for synthesizing the precursor (UDP-glucose is homologous to dual-substrate UDP-galactose/glucose pyrophosphorylases (UGGPases, rather than the "classic" UDP-glucose pyrophosphorylase (UGPase. Phylogenetic recontructions indicate that both T6PS-TPases and UGGPases in apicomplexans and other alveolates are evolutionarily affiliated with stramenopiles and plants. The expression level of T6PS-TPase in C. parvum is highly elevated in the late intracellular developmental stage prior to or during the production of oocysts, implying that trehalose may be important in oocysts as a protectant against environmental stresses. Finally, trehalose has been detected in C. parvum oocysts, thus confirming the trehalose synthetic activity in this parasite. CONCLUSIONS/SIGNIFICANCE: A trehalose synthetic pathway is described in the majority of apicomplexan parasites including Cryptosporidium and the presence of trehalose was confirmed in the C. parvum oocyst. Key enzymes in the pathway (i.e., T6PS-TPase and UGGPase
Kváč, Martin; Hořická, Anna; Sak, Bohumil; Prediger, Jitka; Salát, Jiří; Širmarová, Jana; Bartonička, Tomáš; Clark, Mark; Chelladurai, Jeba Rose Jennifer Jesudoss; Gillam, Erin; McEvoy, John
Bats from the families Rhinolophidae (n = 90) and Vespertilionidae (n = 191) in the USA and Czech Republic were screened for the presence of Cryptosporidium by microscopic and molecular analysis of faecal samples collected from rectum of dissected animals and from the ground beneath roosting sites. Cryptosporidium oocysts were not detected in any of the 281 faecal specimens examined using the aniline-carbol-methyl violet staining method. Nested PCR amplification, sequencing and phylogenetic analysis of the small ribosomal subunit rRNA and actin genes were used to identify isolates and infer evolutionary relationships. Cryptosporidium parvum was identified in a western small-footed bat (Myotis ciliolabrum) from the USA and a common pipistrelle bats (Pipistrellus pipistrellus) from the Czech Republic. Two novel genotypes were identified and named Cryptosporidium bat genotype III and IV. Bat genotype III was found in two big brown bats (Eptesicus fuscus) from the USA. Bat genotype IV was detected in two common pipistrelle bats from the Czech Republic.
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.
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.
E Nazemalhoseini Mojarad
Full Text Available "nBackground: Cryptosporidium is an intracellular apicomplexan parasite that infects a wide range of vertebrates including humans. Cryptosporidiosis is a major cause of diarrhea in children with and without human immunodeficiency virus (HIV infection in developing countries. More recently, the molecular methods for identification of morphologically indistinguishable species have been developed. The aim of this study was to determine the characterization of various species of this coccidian among children with diarrhea by using molecular methods."nMethods: Fecal samples were collected from 1263 children with diarrhea who referred to Pediatrics Medical Centers in Qazvin and Tehran, two central provinces of Iran. Initial identification of Cryptosporidium was carried out by Zeihl-Neelsen acid-fast staining method of stool samples. DNA was extracted from positive microscopically samples and were subjected to a two step nested PCR-RFLP based on SSU-rRNA gene."nResults: Out of 1263 collected samples, 31 (2.5% were found to be contained Cryptosporidium oocysts. RFLP analysis showed that 80.6% of the positive isolates were Cryptosporidium parvum, 16.1% C. hominis and 3.2% had mix infection pattern of both C. parvum and C. hominis."nConclusion: Our results showed that the zoonotic pattern of transmission is predominant and has considerable significance in epidemiology of cryptosporidiosis in the study areas.
Ondriska, František; Vrabcová, Ivana; Brinďáková, Silvia; Kváč, Martin; Ditrich, Oleg; Boldiš, Vojtech; Bastlová, Marcela
Cryptosporidiosis belongs to the important parasitic infections with zoonotic potential and the occurrence in European countries is rare. The first cases of cryptosporidiosis caused by Cryptosporidium hominis detected in the Slovak republic were described here. Collection of examined humans consisted of five family members. Faecal specimens were examined by formalin sedimentation, by the Sheather's sugar flotation and by immunochromatography and visualised by the Ziehl-Neelsen acid fast stain. A fragment of the Cryptosporidium small subunit ribosomal RNA gene was amplified by nested polymerase chain reaction and species was determined by restriction fragment length polymorphism analysis with the endonucleases SspI and VspI. C. hominis was found in faeces of two immunocompetent siblings (a 7-year-old boy and a 2-year-old girl). The symptoms occurred only in the boy as gastrointestinal disorders lasting 5 days, and manifested by abdominal pain, an elevated body temperature (37.2 °C), mild diarrhoea, accompanied by lassitude, depression and anorexia. Ultrasonic scan revealed enlarged spleen and mezenteric lymph nodes. Microscopic examination of the stool sample revealed numerous Cryptosporidium oocysts. The DNA typing identified C. hominis subtype IbA10G2. Cryptosporidium was also detected in the boy's sister without any complications and symptoms. Their father, mother and grandmother were parasitologically negative. The source of infection remained unknown. Human cases in present study reflect necessity of systematic attention on intestinal parasites diagnostic inclusive of cryptosporidia.
Full Text Available This study was planned to determine the role of Cryptosporidium sp. and other intestinal parasites in the diarrheal diseases in children with 0-15 years old Van district.Materials and methods: In this study, stool samples of 450 children were examined for parasites. In the study, nativ-lugol, formaldehyde-ethyl acetate sedimentation methods and trichrome staining methods were used to detect parasites in stool samples. Additionally, sedimentation methods and modified acid fast staining method were used to detect the Cryptosporidium oocysts.Results: Parasites were found in 154 (34.2% among 450 children’s with diarrhea. In this study; the ratios of parasites were as follow: Giardia intestinalis 13.5%, Blastocystis hominis 10%, Entamoeba coli 3.78%, Cryptosporidium spp. 2.2%, Hymenolepis nana 1.33 %ve Ascaris lumbricoides 1.11%.Entamoeba histolytica/Entamoeba dispar 0.89%, Chilomastix mesnili 1.78%, Iodamoeba butschlii 0.89%, Entamoeba hartmanni 0.89%, Trichomonas hominis 0.67%, Enteromonas hominis 0.67%,Conclusion: In the investigate, it was found that Giardia intestinalis and Blastocystis hominis were most prominent agents in children with diarrhea in our vicinity and Cryptosporidium spp also was an important agent which should be investigated carefully in especially risk group in routine laboratory studies.
Jiao, Bing-xin; Wang, Hui-zhu; Liu, Ying; Li, Juan; Guo, Jie; Li, Min; Wan, Gang; Hua, Wen-hao
To investigate the clinical feature of cryptosporidium infection in HIV/AIDS patients with chronic diarrhea. 253 Stool samples were collected from HIV/AIDS patients with chronic diarrhea during Nov.2009 to Dec.2010. The samples were concentrated by Formalin-Ethyl Acetate Sedimentation technique and stained by Modified acid-fast stain (AFS) for the identification of oocysts by microscopy. Divided the cases into three groups according to their CD4 cell counts (AIDS patients was 12.6% in 253 cases. CD4(+) T-lymphocyte counts was related to the infection rates of cryptosporidium, the difference was statistically significant (χ(2) = 10.33, P 0.05). HIV/AIDS patients with chronic diarrhea who progressed during asymptomatic period, pre-AIDS period, AIDS period, had the infection rate of 0(0/7), 21.3% (19/89), 8.3% (13/157) respectively, the difference was statistically significant (χ(2) = 9.822, P 0.05). The infection rate of cryptosporidium and clinical severity of cryptosporidium infection are statistically correlated with CD4(+) T-lymphocyte counts, with AIDS stage, with HIV associated proctitis.
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.
Vanee, Niti; Roberts, Seth B; Fong, Stephen S; Manque, Patricio; Buck, Gregory A
The apicomplexan Cryptosporidium is a protozoan parasite of humans and other mammals. Cryptosporidium species cause acute gastroenteritis and diarrheal disease in healthy humans and animals, and cause life-threatening infection in immunocompromised individuals such as people with AIDS. The parasite has a one-host life cycle and commonly invades intestinal epithelial cells. The current genome annotation of C. hominis, the most serious human pathogen, predicts 3884 genes of which ca. 1581 have predicted functional annotations. Using a combination of bioinformatics analysis, biochemical evidence, and high-throughput data, we have constructed a genome-scale metabolic model of C. hominis. The model is comprised of 213 gene-associated enzymes involved in 540 reactions among the major metabolic pathways and provides a link between the genotype and the phenotype of the organism, making it possible to study and predict behavior based upon genome content. This model was also used to analyze the two life stages of the parasite by integrating the stage-specific proteomic data for oocyst and sporozoite stages. Overall, this model provides a computational framework to systematically study and analyze various functional behaviors of C. hominis with respect to its life cycle and pathogenicity.
Bouzid, Maha; Hunter, Paul R; McDonald, Vincent; Elwin, Kristin; Chalmers, Rachel M; Tyler, Kevin M
Cryptosporidiosis is predominantly caused by two closely related species of protozoan parasites the zoonotic Cryptosporidium parvum and anthroponotic Cryptosporidium hominis which diverge phenotypically in respect to host range and virulence. Using comparative genomics we identified two genes displaying overt heterogeneity between species. Although initial work suggested both were species specific, Cops-1 for C. parvum and Chos-1 for C. hominis, subsequent study identified an abridged ortholog of Cops-1 in C. hominis. Cops-1 and Chos-1 showed limited, but significant, similarity to each other and share common features: (i) telomeric location: Cops-1 is the last gene on chromosome 2, whilst Chos-1 is the first gene on chromosome 5, (ii) encode circa 50-kDa secreted proteins with isoelectric points above 10, (iii) are serine rich, and (iv) contain internal nucleotide repeats. Importantly, Cops-1 sequence contains specific SNPs with good discriminatory power useful epidemiologically. C. parvum-infected patient sera recognized a 50-kDa protein in antigen preparations of C. parvum but not C. hominis, consistent with Cops-1 being antigenic for patients. Interestingly, anti-Cops-1 monoclonal antibody (9E1) stained oocyst content and sporozoite surface of C. parvum only. This study provides a new example of protozoan telomeres as rapidly evolving contingency loci encoding putative virulence factors. PMID:23467513
Full Text Available Entre agosto de 1990 y diciembre de 1991 se examinaron 120 muestras de materia fecal de niños o adultos que consultaron por diarrea, sugestiva de ser causada por Cryptosporidium spp. En todos los casos se realizó la coloración con Lugol-Nigroslna, que proponemos, y se hizo la confirmación con la de Ziehl Neelsen modificada, pese a su limitación de teñir con el mismo patrón de coloración el Cryptosporldium y estructuras diferentes a él. En 20 (16.6% muestras (12 de niños y 8 de adultos se identificaron ooquistes de Cryptosporldlum spp y todas se confirmaron como positivas por la coloración de Ziehl Neelsen modificada. Dado que no siempre es fácil la observación de parásitos de poca prevalencia sugerimos esta coloración como ensayo de rutina porque ayuda a distinguir los ooquistes de Cryptosporidium y mejora la observación de todos los protozoarios.
We examined 120 stool specimens from patients with diarrheal disease, suspected of being infected with Cryptosporidium. Preliminary observation was made with a Lugol-Nigrosine stain and confirmation with modified Ziehl-Neelsen. Twenty specimens (12 from children and 8 from adults (16.6% were positive for Cryptosporidium oocysts andevery one of them was confirmed with ZN stain. Since It may be difficult to detect low-prevalence parasites we suggest routine use of Lugol-Nigrosine which is useful for the detection of Cryptosporidium as well as of other protozoa.
Roberto César Araujo Lima
Full Text Available Cryptosporidiosis is a waterborne disease, has as aggravating the difficulty of preventing environmental contamination and lack of effective therapeutic measures. With marked importance to the cattle, causes inflammation and intestinal villous atrophy resulting in loss of absorptive surface. This study aimed to perform molecular characterization of Cryptosporidium spp. in calves in the city of Formiga, Minas Gerais. A total of 300 faeces samples from Holstein calves, Nelore and indefinite breed, both healthy, were evaluated by negative contrast staining technique of malachite green and through the reaction of nested PCR for amplification of DNA fragments of the 18S subunit of the RNA gene ribosomal. Occurrence of 5.33 % ( 16/300 for malachite green and 4.66 % ( 14/300 by PCR was observed, whereas no correlation was found between positive and variables studied. Through molecular characterization were identified Cryptosporidium andersoni and Cryptosporidium ryanae species. In conclusion, we observed a low incidence of infection and elimination of Cryptosporidium spp. oocysts, the absence of clinical signs in animals, strong agreement between the results obtained by the two techniques. Beyond, with the molecular characterization ( nested PCR , species of C. andersoni and C. ryanae were diagnosed in age groups not present in the literature. These two species of Cryptosporidium are described above for the first time parasitizing cattle in the state of Minas Gerais.
Full Text Available Considering the proximity of sheep farmers to animals that are possibly diseased or releasing fecal oocysts into the environment and the marked pathogenicity in lambs, the aim of this study was to determine the occurrence and to molecularly characterize the infection by Cryptosporidium spp. in lambs in the South Central region of the state of São Paulo, Brazil. A total of 193 fecal samples were collected from sheep of several breeds, males and females, aged up to one year. Polymerase chain reaction (nested-PCR was used to amplify DNA fragments from the subunit 18S rRNA gene and indicated 15% positivity; sequencing of amplified fragments was possible for 19 samples. Analysis of the obtained sequences showed that the identified species were Cryptosporidium xiaoi for 15 samples, constituting thus the first molecular characterization study of this Cryptosporidium species in Brazil. Cryptosporidium ubiquitum was identified for three samples and Cryptosporidium meleagridis for one sample; the latter two are considered zoonotic species.
Aurélien J. Mazurie
Full Text Available Until recently, the apicomplexan parasites, Cryptosporidium hominis and C. parvum, were considered the same species. However, the two parasites, now considered distinct species, exhibit significant differences in host range, infectivity, and pathogenicity, and their sequenced genomes exhibit only 95–97% identity. The availability of the complete genome sequences of these organisms provides the potential to identify the genetic variations that are responsible for the phenotypic differences between the two parasites. We compared the genome organization and structure, gene composition, the metabolic and other pathways, and the local sequence identity between the genes of these two Cryptosporidium species. Our observations show that the phenotypic differences between C. hominis and C. parvum are not due to gross genome rearrangements, structural alterations, gene deletions or insertions, metabolic capabilities, or other obvious genomic alterations. Rather, the results indicate that these genomes exhibit a remarkable structural and compositional conservation and suggest that the phenotypic differences observed are due to subtle variations in the sequences of proteins that act at the interface between the parasite and its host.
Acción de distintos coagulantes para la eliminación de Cryptosporidium spp. en el proceso de potabilización del agua The action of different coagulants to remove Cryptosporidium during the process of water treatment
Full Text Available Cryptosporidium es uno de los microorganismos de mayor interés desde el punto de vista de la Salud Pública y constituye un problema prioritario para las plantas y organismos reguladores de agua. Debido a su pequeño tamaño y a su resistencia a la cloración, la eliminación por el proceso de potabilización es una tarea compleja. En este trabajo se analizó la efectividad de distintos coagulantes utilizados comúnmente en tal proceso para lograr la remoción de los ooquistes. Se trabajó con la prueba de jarras (Jar Test. Se halló que: 1 Los coagulantes con agregado de polímeros coadyuvantes producen remociones de ooquistes superiores a 2 log. 2 Un valor bajo de turbiedad no asegura una remoción óptima de los parásitos. 3 La adición de polielectrolitos al cloruro férrico disminuye la variabilidad tanto en la turbiedad final como en la eliminación de Cryptosporidium.Cryptosporidium is one of the microorganisms of main concern from the point of view of Public Health, being a priority problem for water treatment plants and water regulatory institutions. Due to its small size and resistance to chlorination, Cryptosporidium removal during the process of drinking water treatmentis a hard task. The effectiveness of different coagulants commonly used in the process of removal of oocysts was analyzed. Thetechnique used was the Jar Test. It was found that: 1 coagulants with the addition of polimeric coadjuvants produce over 2 logs of oocyst removal; 2 a low value in turbidity does not necessarily mean optimal parasite removal, and 3 the addition of polyelectrolites to ferric chloride diminishes variability, both in final turbidity and Cryptosporidium removal.
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: firstname.lastname@example.org [MRC Clinical Sciences Centre, Faculty of Medicine, Imperial College, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN (United Kingdom)
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.
Sikora, Per; Andersson, Sofia; Winiecka-Krusnell, Jadwiga; Hallström, Björn; Alsmark, Cecilia; Troell, Karin; Beser, Jessica; Arrighi, Romanico B G
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. Copyright © 2017 American Society for Microbiology.
Fontán-Sainz, María; Gómez-Couso, Hipólito; Fernández-Ibáñez, Pilar; Ares-Mazás, Elvira
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
Petersen, Heidi Huus; Jianmin, Wang; Katakam, Kiran K.
Although pigs are commonly infected with Cryptosporidium spp. and Giardia duodenalis, including potentially zoonotic species or genotypes, little is known about age-related infection levels, seasonal differences and genetic variation in naturally infected pigs raised in organic management systems....... Therefore, the current study was conducted to assess seasonal and age-related variations in prevalence and infection intensity of Cryptosporidium and Giardia, evaluate zoonotic potential and uncover correlations between species/genotypes, infection intensity and faecal consistency. Shedding of oocysts...... and cysts ((oo-) cysts) was monitored at quarterly intervals (September 2011 to June 2012) in piglets (n=152), starter pigs (n=234), fatteners (n=230) and sows (n=240) from three organic farms in Denmark. (Oo-) cysts were quantified by immunofluorescence microscopy; and 56/75 subsamples from Cryptosporidium...
Hijnen, W.A.M.; Dullemont, Y.J.; Bosklopper, K.T.G.J.; Schijven, J.F.; Medema, Gerriet Jan
Decimal Elimination Capacity (DEC) of the slow sand filters of the Dutch drinking water Companies was assessed; first by literature review, followed by evaluation of the removal of environmental spores of sulphite-reducing clostridia (SSRC) and small-sized centric diatoms (SSCD) as surrogates.
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...
Nada Abu Samra
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
João Carlos Gomes Borges
Full Text Available A criptosporidiose constitui-se como uma zoonose que pode afetar o homem e uma ampla variedade de animais domésticos e silvestres, principalmente indivíduos imunodeficientes. O objetivo desse trabalho foi registrar a ocorrência de infecção por Cryptosporidium em peixe-boi marinho. Após ser constatada a mudança de comportamento de um peixe-boi marinho mantido nos oceanários do Centro Mamíferos Aquáticos, ICMBio - FMA, animal foi submetido à exame clínico e, posteriormente, à coleta de amostra fecal. As amostras fecais foram analisadas pela técnica de Kinyoun, teste de imunofluorescência direta e pelo corante 4'.6'-Diamidino-2-Phenilindole (DAPI. No exame clínico, o animal apresentou sinais de desconforto abdominal. Os resultados obtidos nas análises de microscopia de luz e fluorescente revelaram a presença de oocistos de Cryptosporidium nas fezes desse peixe-boi.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.
Full Text Available Giardia, Entamoeba, Isospora and Cryptosporidium are important protozoan parastites that caused diarrhea in human and animals. In the present study, fecal samples were collected fresh, directly from the rectum of 112 stray dogs in Ilam province. Giardia and Entamoeba were concentrated by using the formalin ether sedimentation method followed by the trichrome and iodine staining technique andCryptosporidium oocysts were concentrated by using the formalin ether sedimentation method followed by the modified Ziehl-Neelsen staining technique. Of 112 stray dogs, protozoan infections were detected from feces of 46 dogs (41.07% that Giardia infection was detected from feces of 21 dogs (18.75%, Isospora 17 (15.17%, Cryptosporidium 8 (7.14% and synchronization infection to 2 protozoan in 9 dogs (8.03% and to 3 protozoan in 3 (2.67%. In the present study not observed to Entamoeba. No statistically significant differences in prevalence of protozoan parasites occurred between female (34.21 % and male (55.5 % stray dogs (p>0/05. But statistically significant differences in prevalence occurred between 1≥0 and 0 ≥1 stray dogs (p>0/05. So that stray dogs of Ilam province can cause infection of human water and food sources.
Amburgey, James E; Anderson, J Brian
Cryptosporidium is a chlorine-resistant protozoan parasite responsible for the majority of waterborne disease outbreaks in recreational water venues in the USA. Swim diapers are commonly used by diaper-aged children participating in aquatic activities. This research was intended to evaluate disposable swim diapers for retaining 5-μm diameter polystyrene microspheres, which were used as non-infectious surrogates for Cryptosporidium oocysts. A hot tub recirculating water without a filter was used for this research. The microsphere concentration in the water was monitored at regular intervals following introduction of microspheres inside of a swim diaper while a human subject undertook normal swim/play activities. Microsphere concentrations in the bulk water showed that the majority (50-97%) of Cryptosporidium-sized particles were released from the swim diaper within 1 to 5 min regardless of the swim diaper type or configuration. After only 10 min of play, 77-100% of the microspheres had been released from all swim diapers tested. This research suggests that the swim diapers commonly used by diaper-aged children in swimming pools and other aquatic activities are of limited value in retaining Cryptosporidium-sized particles. Improved swim diaper solutions are necessary to efficiently retain pathogens and effectively safeguard public health in recreational water venues.
Shimelis, Techalew; Tadesse, Endale
Gastro-enteritis is associated with significant morbidity and mortality in patients with HIV/AIDS and children, and Cryptosporidium is the most important parasite implicated. To date, several commercial companies have developed simple and rapid point-of-care tests for the detection of Cryptosporidium infection; however, information is scarce regarding their diagnostic significance in Ethiopia. This study aimed at evaluating the performance of a rapid diagnostic test (RDT) for the detection of Cryptosporidium stool antigen. A hospital-based cross-sectional study was conducted in Hawassa University Hospital, southern Ethiopia from May to November 2013. Faecal samples were collected from a total of 100 children and 250 HIV infected individuals with diarrhea or CD4 T-cell count lower than 200 cells/μl. Specimens were processed using direct, formol-ether concentration and modified Ziehl-Neelsen techniques for diagnosis of Cryptosporidium and other parasites. One hundred faecal samples (50 positives for Cryptosporidium, 35 positives for other parasites and 15 negatives for any intestinal parasites) were tested using the CoproStrip™Cryptosporidium kit (Savyon Diagnostics Ltd, Israel). Test parameters were calculated using microscopy of the modified Ziehl-Neelsen stained stool smear as reference method. The performance of the RDT was first compared to routine microscopic analysis (examination ≤10 min). The CoproStrip™Cryptosporidium RDT correctly detected 31 of 42 positive samples and 49 of 50 negative samples (i.e., 11 false negatives and 1 false positive). Sensitivity, specificity, PPV, NPV and accuracy were calculated to be 74, 98, 97, 84 and 88%, respectively. Upon thorough microscopic analysis (examination >10 min), 8 more samples with very low oocyst density were found. However, these were missed by the kit and lower the sensitivity and NPV to 62 and 72%, respectively. No cross-reactivity was observed with any of the helminthic or other protozoan parasites
Bartelt, Luther A.; Bolick, David T.; Kolling, Glynis L.; Zaenker, Edna I.; Lara, Ana M.; Noronha, Francisco Jose; Cowardin, Carrie A.; Moore, John H.; Turner, Jerrold R.; Warren, Cirle A.; Buck, Gregory A.; Guerrant, Richard L.
Cryptosporidium is a major cause of severe diarrhea, especially in malnourished children. Using a murine model of C. parvum oocyst challenge that recapitulates clinical features of severe cryptosporidiosis during malnutrition, we interrogated the effect of protein malnutrition (PM) on primary and secondary responses to C. parvum challenge, and tested the differential ability of mucosal priming strategies to overcome the PM-induced susceptibility. We determined that while PM fundamentally alters systemic and mucosal primary immune responses to Cryptosporidium, priming with C. parvum (106 oocysts) provides robust protective immunity against re-challenge despite ongoing PM. C. parvum priming restores mucosal Th1-type effectors (CD3+CD8+CD103+ T-cells) and cytokines (IFNγ, and IL12p40) that otherwise decrease with ongoing PM. Vaccination strategies with Cryptosporidium antigens expressed in the S. Typhi vector 908htr, however, do not enhance Th1-type responses to C. parvum challenge during PM, even though vaccination strongly boosts immunity in challenged fully nourished hosts. Remote non-specific exposures to the attenuated S. Typhi vector alone or the TLR9 agonist CpG ODN-1668 can partially attenuate C. parvum severity during PM, but neither as effectively as viable C. parvum priming. We conclude that although PM interferes with basal and vaccine-boosted immune responses to C. parvum, sustained reductions in disease severity are possible through mucosal activators of host defenses, and specifically C. parvum priming can elicit impressively robust Th1-type protective immunity despite ongoing protein malnutrition. These findings add insight into potential correlates of Cryptosporidium immunity and future vaccine strategies in malnourished children. PMID:27467505
Edwards, Hanna; Thompson, R C Andrew; Koh, Wan H; Clode, Peta L
The Apicomplexan parasite Cryptosporidium parvum is responsible for the widespread disease cryptosporidiosis, in both humans and livestock. The nature of C. parvum infection is far from understood and many questions remain in regard to host-parasite interactions, limiting successful treatment of the disease. To definitively identify a range of C. parvum stages in cell culture and to begin to investigate host cell interactions in some of the lesser known life stages, we have utilized a combined scanning electron microscopy and immunolabeling approach, correlating high resolution microstructural information with definitive immunogold labeling of Cryptosporidium stages. Several life cycle stages, including oocysts, merozoites I, trophozoites, gamonts and microgametocytes, were successfully immunolabeled in an in vitro model system. Developing oocysts were clearly immunolabeled, but this did not persist once excystation had occurred. Immunolabeling visualized on the host cell surface adjacent to invasive merozoites is likely to be indicative of receptor shedding, with merozoites also initiating host responses that manifested as abnormal microvilli on the host cell surface. Small sub-micron stages such as microgametocytes, which were impossible to identify as single entities without immunolabeling, were readily visualized and observed to attach to host cells via novel membranous projections. Epicellular parasites also expressed Cryptosporidium-derived epitopes within their encapsulating membrane. These data have allowed us to confidently identify a variety of C. parvum stages in cell culture at high resolution. With this, we provide new insight into C. parvum - host cell interactions and highlight future opportunities for investigating and targeting receptor-mediated interactions between Cryptosporidium life cycle stages and host cells. Copyright © 2011 Elsevier Ltd. All rights reserved.
Utaaker, Kjersti Selstad; Kumar, Anil; Joshi, Himanshu; Chaudhary, Suman; Robertson, Lucy J
Fresh produce has been recognized as a vehicle of infection for protozoan parasites, particularly Cryptosporidium, and, to a lesser extent, Giardia. For both parasites, outbreaks associated with fresh produce have been documented. Although documented outbreaks tend to be from industrialized countries, contamination of fresh produce with these parasites is a global issue. In developing countries, infections with these parasites are often endemic in the community, and basic infrastructure and hygiene measures may be inadequate, thus the likelihood of contamination of fresh produce with these parasites may be higher. Realization of the importance of this transmission route comes against a backdrop of raw salads and more Western culinary habits gaining a foothold, and fresh produce being encouraged as part of the diet due to their associated health benefits. However, if consumption of uncooked fresh produce is going to increase its market sector in India, it is important that it is safe. In this study, various types of fresh produce obtained from three types of vendors in Chandigarh, a major city in Northern India, were analyzed for contamination with Cryptosporidium oocysts and Giardia cysts using a method that has been previously validated in inter-laboratory spiking experiments. A total of 284 samples of different fresh produce items were analyzed, obtained from the different retailers situated in different societal layers of the city. The overall prevalence of contamination of fresh produce with these parasites was just under 11%, with 6% of the vegetables contaminated with Cryptosporidium oocysts, and 5% with Giardia cysts. Contaminated vegetables included turnip, cabbage, carrot, chili, coriander, cucumber, radishes, and tomatoes. Molecular analyses identified contamination with Cryptosporidium parvum and Giardia duodenalis of Assemblage A and Assemblage D, indicating that contamination from animals may be of relevance. Although the prevalence of contamination is
Mateo, Marta; de Mingo, Marta Hernández; de Lucio, Aida; Morales, Lucía; Balseiro, Ana; Espí, Alberto; Barral, Marta; Lima Barbero, José Francisco; Habela, Miguel Ángel; Fernández-García, José L; Bernal, Rafael Calero; Köster, Pamela C; Cardona, Guillermo A; Carmena, David
There is a surprisingly scarce amount of epidemiological and molecular data on the prevalence, frequency, and diversity of the intestinal protozoan parasites Giardia duodenalis and Cryptosporidium spp. in wildlife in general and mesocarnivore species in particular. Consequently, the extent of the cyst/oocyst environmental contamination attributable to these wild host species and their potential implications for public veterinary health remain largely unknown. In this molecular epidemiological survey a total of 193 individual faecal samples from badgers (Meles meles, n=70), ferrets (Mustela putorius furo, n=2), genets (Genetta genetta, n=6), Iberian lynxes (Lynx pardinus, n=6), beech martens (Martes foina, n=8), mongooses (Herpestes ichneumon, n=2), otters (Lutra lutra, n=2), polecats (Mustela putorius, n=2), red foxes (Vulpes vulpes, n=87), wildcats (Felis silvestris, n=2), and wolves (Canis lupus, n=6) were obtained from road-killed, hunted, and accidentally found carcasses, and from camera-trap surveys or animals entering rescue shelters, during the period December 2003-April 2016. Investigated specimens were collected in five Spanish autonomous regions including Andalusia (n=1), Asturias (n=69), Basque Country (n=49), Castile-La Mancha (n=38), and Extremadura (n=36). The presence of cysts/oocysts was confirmed by PCR-based methods targeting the small subunit (ssu) ribosomal RNA gene of these parasite species. Genotyping of the obtained isolates were attempted at appropriate markers including the glutamate dehydrogenase (G. duodenalis) and the 60-kDa glycoprotein (C. parvum and C. ubiquitum) loci. Overall, G. duodenalis was detected in 8% (7/87) of red foxes, a single beech marten, and a single wolf, respectively. Cryptosporidium was identified in 3% (2/70) of badgers, 8% (7/87) of red foxes, a single genet, and a single mongoose, respectively. None of the nine G. duodenalis isolates generated could be genotyped at the assemblage/sub-assemblage level. Out of the
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.
Metge, D W; Harvey, R W; Aiken, G R; Anders, R; Lincoln, G; Jasperse, J
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 microm) 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 (D(50)) 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 microM) 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. Published by Elsevier Ltd.
Ch Stratakos, Alexandros; Sima, Filip; Ward, Patrick; Linton, Mark; Kelly, Carmel; Pinkerton, Laurette; Stef, Lavinia; Pet, Ioan; Iancu, Tiberiu; Pircalabioru, Gratiela; Corcionivoschi, Nicolae
Cryptosporidium is a major cause of diarrhea worldwide in both humans and farm animals with no completely effective treatment available at present. In this study, we assessed the inhibitory effect of different concentrations of Auranta 3001 (0.1, 0.5 and 1%), a novel natural feed supplement, on C. hominis and C. parvum invasion of human ileocecal adenocarcinoma (HCT-8), bovine primary cells and C. parvum invasion of HCT-8, bovine primary cells and bovine intestinal biopsies. The effect of the feed supplement on the production of pro-inflammatory cytokines IL-8 and INF-γ, the anti-inflammatory cytokine IL-10, the expression of CpSUB1 protease gene during infection was also assessed by quantitative PCR (q-PCR). Transepithelial electrical resistance (TEER) was employed to measure the integrity of tight junction dynamics of the culture models. Pre-treatment of intestinal cells or oocysts with the Auranta 3001 significantly reduced the invasiveness of C. hominis and C. parvum against HCT-8 and bovine primary cells in a dose dependent manner. The most pronounced reduction in the invasiveness of both parasites was observed when Auranta 3001 was present during infection. Levels of IL-8 were significantly reduced in both HCT-8 and bovine primary cells, while the levels of INF-γ and IL-10 showed opposite trends in the two cell lines during infection in the presence of Auranta 3001. CpSUB1 gene protease expression, which mediates infection, was significantly reduced suggesting that this enzyme is a possible target of Auranta 3001. Although, C. hominis and C. parvum use different invasion mechanisms to infect cells, the novel feed additive can significantly attenuate the entry of Cryptosporidium in HCT-8 cells, primary bovine cells and bovine intestinal biopsies and thus provide an alternative method to control cryptosporidiosis.
... (WHO) estimates of illness and mortality and epidemiological investigations in the United States still indicates that the threat to public health from waterborne disease still exists There are a plethora of contaminants...
Oocysts are important stage for the spread of Toxoplasma gondii because they are environmentally resistant. Among all hosts of T. gondii, only felids can excrete oocysts. Cats that have excreted T. gondii oocysts after primary infection become immune to re-excretion of oocysts, and this immunity app...
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
Patrícia de Lucca
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.
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
Full Text Available Cryptosporidium is a leading cause of childhood diarrhea in low-resource settings, and has been repeatedly associated with impaired physical and cognitive development. In May 2013, an outbreak of diarrhea caused by Cryptosporidium hominis was identified in the Arctic region of Nunavik, Quebec. Human cryptosporidiosis transmission was previously unknown in this region, and very few previous studies have reported it elsewhere in the Arctic. We report clinical, molecular, and epidemiologic details of a multi-village Cryptosporidium outbreak in the Canadian Arctic.We investigated the occurrence of cryptosporidiosis using a descriptive study of cases with onset between April 2013 and April 2014. Cases were defined as Nunavik inhabitants of any age presenting with diarrhea of any duration, in whom Cryptosporidium oocysts were detected by stool microscopy in a specialised reference laboratory. Cryptosporidium was identified in stool from 51 of 283 individuals. The overall annual incidence rate (IR was 420 / 100,000 inhabitants. The IR was highest among children aged less than 5 years (1290 /100,000 persons. Genetic subtyping for stool specimens from 14/51 cases was determined by DNA sequence analysis of the 60 kDa glycoprotein (gp60 gene. Sequences aligned with C. hominis subtype Id in all cases. No common food or water source of infection was identified.In this first observed outbreak of human cryptosporidiosis in this Arctic region, the high IR seen is cause for concern about the possible long-term effects on growth and development of children in Inuit communities, who face myriad other challenges such as overcrowding and food-insecurity. The temporal and geographic distribution of cases, as well as the identification of C. hominis subtype Id, suggest anthroponotic rather than zoonotic transmission. Barriers to timely diagnosis delayed the recognition of human cryptosporidiosis in this remote setting.
Thivierge, Karine; Iqbal, Asma; Dixon, Brent; Dion, Réjean; Levesque, Benoît; Cantin, Philippe; Cédilotte, Lyne; Ndao, Momar; Proulx, Jean-François; Yansouni, Cedric P
Cryptosporidium is a leading cause of childhood diarrhea in low-resource settings, and has been repeatedly associated with impaired physical and cognitive development. In May 2013, an outbreak of diarrhea caused by Cryptosporidium hominis was identified in the Arctic region of Nunavik, Quebec. Human cryptosporidiosis transmission was previously unknown in this region, and very few previous studies have reported it elsewhere in the Arctic. We report clinical, molecular, and epidemiologic details of a multi-village Cryptosporidium outbreak in the Canadian Arctic. We investigated the occurrence of cryptosporidiosis using a descriptive study of cases with onset between April 2013 and April 2014. Cases were defined as Nunavik inhabitants of any age presenting with diarrhea of any duration, in whom Cryptosporidium oocysts were detected by stool microscopy in a specialised reference laboratory. Cryptosporidium was identified in stool from 51 of 283 individuals. The overall annual incidence rate (IR) was 420 / 100,000 inhabitants. The IR was highest among children aged less than 5 years (1290 /100,000 persons). Genetic subtyping for stool specimens from 14/51 cases was determined by DNA sequence analysis of the 60 kDa glycoprotein (gp60) gene. Sequences aligned with C. hominis subtype Id in all cases. No common food or water source of infection was identified. In this first observed outbreak of human cryptosporidiosis in this Arctic region, the high IR seen is cause for concern about the possible long-term effects on growth and development of children in Inuit communities, who face myriad other challenges such as overcrowding and food-insecurity. The temporal and geographic distribution of cases, as well as the identification of C. hominis subtype Id, suggest anthroponotic rather than zoonotic transmission. Barriers to timely diagnosis delayed the recognition of human cryptosporidiosis in this remote setting.
Cryptosporidium species and Giardia intestinalis cause diarrheal infections in humans and other vertebrate animals globally and are considered to be of great public health importance. The study was conducted to determine the prevalence Cryptosporidium species and G. intestinalis infections among patients attending ...
This study was carried out to assess the potential of animals, used for teaching and research, as a source of Cryptosporidium infection for students and staff of a University in Nigeria. Faecal samples from 185 animals reared on the teaching and research farm were collected and examined for Cryptosporidium spp. antigens ...
Hofstra, N.; Bouwman, A.F.; Beusen, A.H.W.; Medema, G.J.
The protozoan parasite Cryptosporidium is a major cause of diarrhoea worldwide. This paper presents the first model-based inventory with 0.5 by 0.5 degree resolution of global Cryptosporidium emissions for the year 2000 from humans and animals to surface water. The model is based on nutrient
Xiao, Lihua; Ryan, Una M.; Graczyk, Thaddeus K.; Limor, Josef; Li, Lixia; Kombert, Mark; Junge, Randy; Sulaiman, Irshad M.; Zhou, Ling; Arrowood, Michael J.; Koudela, Břetislav; Modrý, David; Lal, Altaf A.
The genetic diversity of Cryptosporidium in reptiles was analyzed by PCR-restriction fragment length polymorphism and sequence analysis of the small subunit rRNA gene. A total of 123 samples were analyzed, of which 48 snake samples, 24 lizard samples, and 3 tortoise samples were positive for Cryptosporidium. Nine different types of Cryptosporidium were found, including Cryptosporidium serpentis, Cryptosporidium desert monitor genotype, Cryptosporidium muris, Cryptosporidium parvum bovine and mouse genotypes, one C. serpentis-like parasite in a lizard, two new Cryptosporidium spp. in snakes, and one new Cryptosporidium sp. in tortoises. C. serpentis and the desert monitor genotype were the most common parasites and were found in both snakes and lizards, whereas the C. muris and C. parvum parasites detected were probably the result of ingestion of infected rodents. Sequence and biologic characterizations indicated that the desert monitor genotype was Cryptosporidium saurophilum. Two host-adapted C. serpentis genotypes were found in snakes and lizards. PMID:14766569
Timothy S. Mamba
Full Text Available Background. Cryptosporidium is a protozoan parasite and a major cause of diarrhea in children and immunocompromised patients. Current diagnostic methods for cryptosporidiosis such as microscopy have low sensitivity while techniques such as PCR indicate higher sensitivity levels but are seldom used in developing countries due to their associated cost. A loop-mediated isothermal amplification (LAMP technique, a method with shorter time to result and with equal or higher sensitivity compared to PCR, has been developed and applied in the detection of Cryptosporidium species. The test has a detection limit of 10 pg/µl (~100 oocysts/ml indicating a need for more sensitive diagnostic tools. This study developed a more sensitive lateral flow dipstick (LFD LAMP test based on SAM-1 gene and with the addition of a second set of reaction accelerating primers (stem primers. Results. The stem LFD LAMP test showed analytical sensitivity of 10 oocysts/ml compared to 100 oocysts/ml (10 pg/ul for each of the SAM-1 LAMP test and nested PCR. The stem LFD LAMP and nested PCR detected 29/39 and 25/39 positive samples of previously identified C. parvum and C. hominis DNA, respectively. The SAM-1 LAMP detected 27/39. On detection of Cryptosporidium DNA in 67 clinical samples, the stem LFD LAMP detected 16 samples and SAM-2 LAMP 14 and nested PCR identified 11. Preheating the templates increased detection by stem LFD LAMP to 19 samples. Time to results from master mix preparation step took ~80 minutes. The test was specific, and no cross-amplification was recorded with nontarget DNA. Conclusion. The developed stem LFD LAMP test is an appropriate method for the detection of C. hominis, C. parvum, and C. meleagridis DNA in human stool samples. It can be used in algorithm with other diagnostic tests and may offer promise as an effective diagnostic tool in the control of cryptosporidiosis.
Ifeonu, Olukemi O; Chibucos, Marcus C; Orvis, Joshua; Su, Qi; Elwin, Kristin; Guo, Fengguang; Zhang, Haili; Xiao, Lihua; Sun, Mingfei; Chalmers, Rachel M; Fraser, Claire M; Zhu, Guan; Kissinger, Jessica C; Widmer, Giovanni; Silva, Joana C
Human cryptosporidiosis is caused primarily by Cryptosporidium hominis, C. parvum and C. meleagridis. To accelerate research on parasites in the genus Cryptosporidium, we generated annotated, draft genome sequences of human C. hominis isolates TU502_2012 and UKH1, C. meleagridis UKMEL1, also isolated from a human patient, and the avian parasite C. baileyi TAMU-09Q1. The annotation of the genome sequences relied in part on RNAseq data generated from the oocyst stage of both C. hominis and C. baileyi The genome assembly of C. hominis is significantly more complete and less fragmented than that available previously, which enabled the generation of a much-improved gene set for this species, with an increase in average gene length of 500 bp relative to the protein-encoding genes in the 2004 C. hominis annotation. Our results reveal that the genomes of C. hominis and C. parvum are very similar in both gene density and average gene length. These data should prove a valuable resource for the Cryptosporidium research community. © FEMS 2016.
Prevalence and genetic characterization of Cryptosporidium spp. and Cystoisospora belli in HIV-infected patients Prevalência e caracterização genética de Cryptosporidium spp. e Cystoisospora belli em pacientes infectados pelo HIV
Dnieber Chagas Assis
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 Cryptosporidium spp. e Cystoisospora belli são protozoários monoxenos reconhecidos como agentes causadores de diarréia crônica em indivíduos imunocomprometidos, especialmente aqueles infectados pelo HIV. Os objetivos deste estudo foram o de avaliar a frequência destes protozoários em pacientes HIV - positivos na região do Triângulo Mineiro, Brasil, e correlacionar a presença destas infecções com dados clínicos, epidemiológicos e laboratoriais dos pacientes. Oocistos foram detectados em amostras fecais de 10 (16,9% dos 59 pacientes estudados, sendo 10.1% (6/59 das amostras positivas para Cryptosporidium spp. e 6,7% (4/59 das amostras positivas para C. belli. A frequência destes parasitos foi maior entre pacientes com síndrome diarreica e contagem de linfócitos T CD4+ < 200 cells/mm 3 , o que demonstra o caráter oportunista destas infecções. Foi observada uma associação significativa entre a falta de aderência à terapia antiretroviral e a presença de Cryptosporidium spp. e/ou C. belli. Parasitismo por Cryptosporidium spp. foi mais frequente em fevereiro e abril, meses subsequentes ao período chuvoso. O mesmo não foi observado para C. belli. A caracterização genética de dois
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
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
H. E. Laubach
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.
Balderrama-Carmona, Ana Paola; Gortáres-Moroyoqui, Pablo; Álvarez-Valencia, Luis Humberto; Castro-Espinoza, Luciano; Balderas-Cortés, José de Jesús; Mondaca-Fernández, Iram; Chaidez-Quiroz, Cristóbal; Meza-Montenegro, María Mercedes
Cryptosporidium and Giardia are gastrointestinal disease-causing organisms transmitted by the fecal-oral route, zoonotic and prevalent in all socioeconomic segments with greater emphasis in rural communities. The goal of this study was to assess the risk of cryptosporidiosis and giardiasis of Potam dwellers consuming drinking water from communal well water. To achieve the goal, quantitative microbial risk assessment (QMRA) was carried out as follows: (a) identification of Cryptosporidium oocysts and Giardia cysts in well water samples by information collection rule method, (b) assessment of exposure to healthy Potam residents, (c) dose-response modelling, and (d) risk characterization using an exponential model. All well water samples tested were positive for Cryptosporidium and Giardia. The QMRA results indicate a mean of annual risks of 99:100 (0.99) for cryptosporidiosis and 1:1 (1.0) for giardiasis. The outcome of the present study may drive decision-makers to establish an educational and treatment program to reduce the incidence of parasite-borne intestinal infection in the Potam community, and to conduct risk analysis programs in other similar rural communities in Mexico.
Freitas, Dayana Andrade de; Paiva, Anderson Luiz Ribeiro de; Carvalho Filho, José Adson Andrade de; Cabral, Jaime Joaquim da Silva Pereira; Rocha, Francisca Janaína Soares
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. 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. Cryptosporidium spp., Giardia spp. and other parasites were found in the Beberibe River. Sanitation companies must assess pathogenic intestinal parasites in water basins providing public water and subsequently develop improved treatment systems for removal of such parasites.
Dayana Andrade de Freitas
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.
Alyousefi, N A; Mahdy, M A K; Lim, Y A L; Xiao, L; Mahmud, R
Cryptosporidium is a protozoan parasite of humans and animals and has a worldwide distribution. The parasite has a unique epidemiology in Middle Eastern countries where the IId subtype family of Cryptosporidium parvum dominates. However, there has been no information on Cryptosporidium species in Yemen. Thus, this study was conducted in Yemen to examine the distribution of Cryptosporidium species and subtype families. Fecal samples were collected from 335 patients who attended hospitals in Sana'a city. Cryptosporidium species were determined by PCR and sequence analysis of the 18 s rRNA gene. Cryptosporidium parvum and C. hominis subtypes were identified based on sequence analysis of the 60 kDa glycoprotein (gp60) gene. Out of 335 samples, 33 (9.9%) were positive for Cryptosporidium. Of them, 97% were identified as C. parvum whilst 1 case (3%) was caused by C. hominis. All 7 C. parvum isolates subtyped belonged to the IIaA15G2R1 subtype. The common occurrence of the zoonotic IIa subtype family of C. parvum highlights the potential occurrence of zoonotic transmission of cryptosporidiosis in Yemen. However, this postulation needs confirmation with future molecular epidemiological studies of cryptosporidiosis in both humans and animals in Yemen.
Debenham, John J; Atencia, Rebeca; Midtgaard, Fred; Robertson, Lucy J
The aim of this study was to investigate the occurrence of Giardia duodenalis and Cryptosporidium spp. in primates and determine their zoonotic or anthropozoonotic potential. Direct immunofluorescence was used to identify Giardia and Cryptosporidium from faecal samples. PCR and DNA sequencing was performed on positive results. Giardia cysts were identified from 5.5% (5/90) of captive chimpanzees and 0% (0/11) of captive mandrills in the Republic of Congo; 0% (0/10) of captive chimpanzees in Norway; and 0% of faecal samples (n = 49) from wild Zanzibar red colobus monkeys. Two Giardia positive samples were also positive on PCR, and sequencing revealed identical isolates of Assemblage B. Cryptosporidium oocysts were not detected in any of the samples. In these primate groups, in which interactions with humans and human environments are quite substantial, Giardia and Cryptosporidium are rare pathogens. In chimpanzees, Giardia may have a zoonotic or anthropozoonotic potential. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.
Ruecker, Norma J.; Hoffman, Rebecca M.; Chalmers, Rachel M.; Neumann, Norman F.
Molecular methods incorporating nested PCR-restriction fragment length polymorphism (RFLP) analysis of the 18S rRNA gene of Cryptosporidium species were validated to assess performance based on limit of detection (LoD) and for detecting and resolving mixtures of species and genotypes within a single sample. The 95% LoD was determined for seven species (Cryptosporidium hominis, C. parvum, C. felis, C. meleagridis, C. ubiquitum, C. muris, and C. andersoni) and ranged from 7 to 11 plasmid template copies with overlapping 95% confidence limits. The LoD values for genomic DNA from oocysts on microscope slides were 7 and 10 template copies for C. andersoni and C. parvum, respectively. The repetitive nested PCR-RFLP slide protocol had an LoD of 4 oocysts per slide. When templates of two species were mixed in equal ratios in the nested PCR-RFLP reaction mixture, there was no amplification bias toward one species over another. At high ratios of template mixtures (>1:10), there was a reduction or loss of detection of the less abundant species by RFLP analysis, most likely due to heteroduplex formation in the later cycles of the PCR. Replicate nested PCR was successful at resolving many mixtures of Cryptosporidium at template concentrations near or below the LoD. The cloning of nested PCR products resulted in 17% of the cloned sequences being recombinants of the two original templates. Limiting-dilution nested PCR followed by the sequencing of PCR products resulted in no sequence anomalies, suggesting that this method is an effective and accurate way to study the species diversity of Cryptosporidium, particularly for environmental water samples, in which mixtures of parasites are common. PMID:21498746
Hofstra, Nynke; Vermeulen, Lucie C
Cryptosporidium is a pathogenic protozoan parasite and is a leading cause of diarrhoea worldwide. The concentration of Cryptosporidium in the surface water is a determinant for probability of exposure and the risk of disease. Surface water concentrations are expected to change with population growth, urbanisation and changes in sanitation. The objective of this paper is to assess the importance of future changes in population, urbanisation and sanitation on global human emissions of Cryptosporidium to surface water. The GloWPa-Crypto H1 (the Global Waterborne Pathogen model for Human Cryptosporidium emissions version 1) model is presented and run for 2010 and with scenarios for 2050. The new scenarios are based on the Shared Socio-economic Pathways (SSPs) developed for the climate community. The scenarios comprise assumptions on sanitation changes in line with the storylines and population and urbanisation changes from the SSPs. In SSP1 population growth is limited, urbanisation large and sanitation and waste water treatment strongly improve. SSP1* is the same as SSP1, but waste water treatment does not improve. SSP3 sees large population growth, moderate urbanisation and sanitation and waste water treatment fractions that are the same as in 2010. Total global Cryptosporidium emissions to surface water for 2010 are estimated to be 1.6×10 17 oocysts per year, with hotspots in the most urbanised parts of the world. In 2050 emissions are expected to decrease by 24% or increase by 52% and 70% for SSP1, SSP3 and SSP1* respectively. The emissions increase in all scenarios for countries in the Middle East and Africa (MAF) region, while emissions in large parts in Europe decrease in scenarios SSP1 and SSP3. Improving sanitation by connecting the population to sewers, should be combined with waste water treatment, otherwise (SSP1*) emissions in 2050 are expected to be much larger than in a situation with strong population growth and slow development of safe water and
Chieffi Pedro Paulo
Full Text Available The frequency of infection by Cryptosporidium parvum was determined in two groups of renal patients submitted to immunosuppression. One group consisted of 23 renal transplanted individuals, and the other consisted of 32 patients with chronic renal insufficiency, periodically submitted to hemodialysis. A third group of 27 patients with systemic arterial hypertension, not immunosuppressed, was used as control. During a period of 18 months all the patients were submitted to faecal examination to detect C. parvum oocysts, for a total of 1 to 6 tests per patient. The results showed frequencies of C. parvum infection of 34.8%, 25% and 17.4%, respectively, for the renal transplanted group, the patients submitted to hemodialysis and the control group. Statistical analysis showed no significant differences among the three groups even though the frequency of C. parvum infection was higher in the transplanted group. However, when the number of fecal samples containing C. parvum oocysts was taken in account, a significantly higher frequency was found in the renal transplanted group.
Toxoplasma gondii oocysts are highly resistant to many chemical sanitizers. Current methods used to determine oocyst infectivity have relied exclusively on mouse, chicken, and feline bioassays. Although considered gold standards, they only provide a qualitative assessment of oocyst infectivity. I...
Stensvold, Christen Rune; Ethelberg, Steen; Hansen, Louise
. RESULTS: A total of 689 Cryptosporidium-positive stool samples were submitted by 387 patients. Limiting case episodes to two months (60 days), a total of 388 case episodes representing 387 patients were identified. Cryptosporidiosis was most common among infants and toddlers. Moreover, a peak in incidence...... was observed among younger adults aged 23-24 years. In 43 Cryptosporidium-positive faecal samples, identification was performed to species and subtype level. Cryptosporidium parvum was found in 34 samples, C. hominis in eight, and C. meleagridis in one sample; C. parvum subtypes IIaA15G2R1 (n = 10) and IIaA16G...
Jarvie, B D; Trotz-Williams, L A; McKnight, D R; Leslie, K E; Wallace, M M; Todd, C G; Sharpe, P H; Peregrine, A S
Thirty-one Holstein bull calves were purchased at birth from 3 dairy farms in Eastern Ontario. Each calf was assigned at random to oral treatment with either 5 mg of halofuginone lactate in 10.0 mL of aqueous carrier solution (Halocur, base comprised 10 mg of benzoic acid, 100 mg of lactic acid, and 0.3 mg of tartrazine) or 10 mL of placebo (Halocur base minus the active ingredient, halofuginone lactate) administered 15 to 30 min after morning milk feeding for the first 7 d of life. Intakes of milk, calf starter, and water, and fecal consistency score were recorded daily for 56 d. Calf weights were recorded weekly for 56 d. Fecal samples were taken from all calves at approximately 2, 7, 14, 21, and 28 d of age for isolation of Cryptosporidium parvum oocysts. Logistic and linear regression analyses were used to assess the effect of treatment on the incidence of diarrhea and C. parvum infection status. The odds of C. parvum shedding among calves in the halofuginone lactate-treated group was 70% lower than the odds of shedding among calves in the placebo group. In calves treated with halofuginone lactate, no oocyst shedding occurred until 2 wk of age, whereas 12.5% of calves in the placebo group began shedding oocysts during wk 1. From all ages of placebo-treated calves, 31 of 73 samples (42.5%) were positive for C. parvum, whereas only 15 of 67 samples (22.4%) from all ages of halofuginone lactate-treated calves tested positive. The largest number of C. parvum-positive samples occurred in the third week of life. There was a significant delay of 3.1 d in the incidence of diarrhea among calves treated with halofuginone lactate. Intake of milk and starter, body weight gains, and age at weaning were not significantly different between treatment groups.
Schets FM; Engels GB; Leenen EJTM; MGB
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
Ethelberg, S.; Lisby, M.; Vestergaard, L. S.
Foodborne outbreaks of cryptosporidiosis are uncommon. In Denmark human cases are generally infrequently diagnosed. In 2005 an outbreak of diarrhoea affected company employees near Copenhagen. In all 99 employees were reported ill; 13 were positive for Cryptosporidium hominis infection. Two...
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Gertler, Maximilian; Dürr, Matthias; Renner, Peter; Poppert, Sven; Askar, Mona; Breidenbach, Janina; Frank, Christina; Preußel, Karina; Schielke, Anika; Werber, Dirk; Chalmers, Rachel; Robinson, Guy; Feuerpfeil, Irmgard; Tannich, Egbert; Gröger, Christine; Stark, Klaus; Wilking, Hendrik
During weeks 32-33, 2013, 24 cases of cryptosporidiosis were notified in the city of Halle (annual mean 2008-2012: 9 cases). We investigated the outbreak to identify the source and recommend control measures, considering that between weeks 23-25 the river Saale which flows through the city centre overflowed the floodplain, parts of the city centre and damaged sewage systems. We defined a case as a resident of Halle with gastroenteritis, Cryptosporidium-positive stool and disease onset weeks 27 through 47. In a case-control study among kindergarten children, we compared cases and controls regarding environmental exposure, use of swimming pools, zoo visits and tap water consumption 14 days pre-onset or a corresponding 14-days-period (controls) and adjusted for residence. Stool specimens were tested by microscopy and PCR, and Cryptosporidium DNA was sequenced. Samples from public water system, swimming pools and river Saale were examined for Cryptosporidium oocysts (microscopy and PCR). Overall, 167 cases were detected, 40/167 (24%) were classified as secondary cases. First disease onsets occurred during week 29, numbers peaked in week 34 and started to decrease in week 36. Median age was 8 years (range: 0-77). Compared to controls (n = 61), cases (n = 20) were more likely to report visits to previously flooded areas (OR: 4.9; 95%-CI: 1.4-18) and the zoo (OR: 2.6; 95%-CI: 0.9-7.6). In multivariable analysis visits to the floodplain remained the sole risk factor (OR: 5.5; 95%-CI: 1.4-22). Only C.hominis of a single genotype (IbA9G2) was detected in stools. Oocysts were detected in samples from the river, two local lakes and three public swimming pools by microscopy, but not in the public water supply. Evidence suggests that activities in the dried out floodplain led to infection among children. Secondary transmissions may be involved. Consequently, authorities recommended to avoid playing, swimming and having picnics in the flood-affected area. Health
Ziehl Neelsen technique modified with Dimetil Sulfóxido (DMSO, Ziehl Neelsen modifed by Henriksen and Pohlens (HP, Ziehl Neelsen (ZN and Kinyoun (K, completing 1872 printings analyzed in optical microscopy (1000 x. At this final number, at the different colorations done, oocysts of the parasite Cryptosporidium sp. were viewed in 18 trachea printings, 42 of bursa de Fabricius and 29 of intestine, resulting positive, 89 printings. From these 89, 44 were identified through the technique DMSO, 32 through HP, three through ZN and 10 through K. Cryptosporidium sp. parasites oocysts were found with more frequency from printings of bursa the Fabricius. The coloration method, of those utilized, was the one which had the better visualization of the oocysts was the DMSO.
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.
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
The surface properties of aged (stored for 10 years) and fresh (recently excreted) oocysts of Toxoplasma gondii were investigated using monoclonal antibody (mAb) and lectin-binding assays. Fresh oocysts bound a wall-specific mAb labeled with fluorescein isothiocyanate while aged oocysts did not. In ...
Nazemalhosseini-Mojarad, Ehsan; Haghighi, Ali; Taghipour, Niloofar; Keshavarz, Akbar; Mohebi, Seyed Reza; Zali, Mohammad Reza; Xiao, Lihua
Cryptosporidium is an intestinal parasite associated with severe acute diarrhea in humans and animals. To investigate subtypes of Cryptosporidium spp. isolated from humans and cattle in Iran, 47 Cryptosporidium parvum (22 from children and 25 from cattle) and three Cryptosporidium hominis from children were characterized by sequence analysis of the 60 kDa glycoprotein (gp60) gene. Nine subtypes (two of C. hominis and seven of C. parvum) in four subtype families were identified. Cattle were mainly infected with C. parvum IIa subtypes and humans mostly with the C. parvum IIa and IId subtypes. Consequently, cattle could be a source of human infection with C. parvum IIa in Iran. However, the occurrence of subtype IId families in Iranian children, suggests that other infection sources might also be involved in C. parvum transmission. To our knowledge, this is the first published record and description of Cryptosporidium subtypes in Iran. Copyright © 2011 Elsevier B.V. All rights reserved.
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.
Jiang, Yanyan; Ren, Jinhua; Yuan, Zhongying; Liu, Aiqin; Zhao, Hong; Liu, Hua; Chu, Lei; Pan, Wei; Cao, Jianping; Lin, Yijin; Shen, Yujuan
Cryptosporidium hominis and C. parvum are usually considered to be the major pathogens responsible for human cryptosporidiosis. However, there have been few studies regarding the molecular epidemiology of Cryptosporidium in human infections in China. Here we investigated Cryptosporidium infection in patients with diarrhea, in Danyang Hospital of Jiangsu Province, China, at the genotype level. A total of 232 stool specimens were collected from outpatients with diarrhea in Danyang Hospital of Jiangsu Province, China, from February 2012 to January 2013. Each specimen was stained from direct fecal smears and examined for Cryptosporidium using modified acid fast staining and microscopy. Moreover, genomic DNA of each fecal sample was screened for the presence of Cryptosporidium with nested PCR, which was genotyped by analyzing the DNA sequences of small subunit rRNA (SSU rRNA). The average infection rate of Cryptosporidium was 1.3% (3/232) by microscopy and subjected to PCR amplification of the SSU rRNA gene of Cryptosporidium, with 9.91% (23/232) being positive for Cryptosporidium with a significant peak in autumn. Based on the SSU rRNA gene, two Cryptosporidium spp. were identified, including C. andersoni (n =21) and C. hominis (n =2). Two types of C. andersoni, designated as A370 + and A370 - , were found in the SSU rRNA gene in our present study, which was 100% homologous to C. andersoni infections derived from dairy calves and goats, respectively. The clinical questionnaires showed no significant difference in age, gender and frequency of diarrhea, but duration of diarrhea was shorter for C. andersoni than that of C. hominis (mean, 2 vs. 4 days; p hominis indicated that different Cryptosporidium species might cause different clinical manifestations.
Wells, Beth; Shaw, Hannah; Hotchkiss, Emily; Gilray, Janice; Ayton, Remedios; Green, James; Katzer, Frank; Wells, Andrew; Innes, Elisabeth
The apicomplexan parasite Cryptosporidium represents a threat to water quality and public health. An important zoonotic species involved in human cryptosporidiosis from contaminated water is Cryptosporidium parvum (C. parvum), the main reservoirs of which are known to be farm livestock particularly neonatal calves, although adult cattle, sheep, lambs and wildlife are also known to contribute to catchment loading of C. parvum. This study aimed to establish Cryptosporidium prevalence, species and genotype in livestock, deer and water in a catchment with a history of Cryptosporidium contamination in the public water supply. A novel method of processing adult ruminant faecal sample was used to concentrate oocysts, followed by a nested species specific multiplex (nssm) PCR, targeting the 18S rRNA gene, to speciate Cryptosporidium. A multilocus fragment typing (MLFT) tool was used, in addition to GP60 sequencing, to genotype C. parvum positive samples. A very high prevalence of Cryptosporidium was detected, with speciation identifying a predominance of C. parvum in livestock, deer and water samples. Four GP60 subtypes were detected within C. parvum with the majority IIaA15G2R1 which was detected in all host species and on all farms. Multilocus fragment typing further differentiated these into 6 highly related multilocus genotypes. The high prevalence of Cryptosporidium detected was possibly due to a combination of the newly developed sample processing technique used and a reflection of the high rates of the parasite present in this catchment. The predominance of C. parvum in livestock and deer sampled in this study suggested that they represented a significant risk to water quality and public health. Genotyping results suggested that the parasite is being transmitted locally within the study area, possibly via free-roaming sheep and deer. Further studies are needed to verify particular host associations with subtypes/MLGs. Land and livestock management solutions to
Martins, Felippe Danyel Cardoso; Ferreira, Fernanda Pinto; de Almeida, Jonatas Campos; Ogawa, Liza; dos Santos, Hannah Lia Ettiene Peruch Lemos; dos Santos, Maíra Moreira; Pinheiro, Filipe Aguera; Navarro, Italmar Teodorico; Garcia, João Luis; Freire, Roberta Lemos
The aims of this study were to verify the prevalence of Cryptosporidium spp. and Giardia spp. in animal feces and drinking water on dairy farms and to identify a possible relation between the exposure factors and the presence of these parasites. Fecal samples from cattle and humans and water samples were collected on dairy farms in Paraná, Brazil. Analysis of (oo)cysts in the feces was performed by the modified Ziehl-Neelsen staining and centrifugal flotation in zinc sulfate. Test-positive samples were subjected to nested PCR amplification of the 18SSU ribosomal RNA gene for identification of Cryptosporidium and Giardia and of the gp60 gene for subtyping of Cryptosporidium. Microbiological analysis of water was carried out by the multiple-tube method and by means of a chromogenic substrate, and parasitological analysis was performed on 31 samples by direct immunofluorescence and nested PCR of the genes mentioned above. Identification of the species of Cryptosporidium was performed by sequencing and PCR with analysis of restriction fragment length polymorphisms. The prevalence of Giardia and Cryptosporidium was higher in calves than in adults. Among the samples of cattle feces, Cryptosporidium parvum was identified in 41 (64%), C. ryanae in eight (12.5%), C. bovis in four (6.3%), C. andersoni in five (7.8%), and a mixed infection in 20 samples (31.3%). These parasites were not identified in the samples of human feces. Thermotolerant coliform bacteria were identified in 25 samples of water (45.5%). Giardia duodenalis and C. parvum were identified in three water samples. The gp60 gene analysis of C. parvum isolates revealed the presence of two strains (IIaA20G1R1 and IIaA17G2R2) in the fecal samples and one (IIaA17G2R1) in the water samples. The presence of coliforms was associated with the water source, structure and degradation of springs, rain, and turbidity. The prevalence of protozoa was higher in calves up to six months of age. C. parvum and G. duodenalis were
Roberta Dos Santos Toledo
Full Text Available The aims of this study were to verify the prevalence of Cryptosporidium spp. and Giardia spp. in animal feces and drinking water on dairy farms and to identify a possible relation between the exposure factors and the presence of these parasites. Fecal samples from cattle and humans and water samples were collected on dairy farms in Paraná, Brazil. Analysis of (oocysts in the feces was performed by the modified Ziehl-Neelsen staining and centrifugal flotation in zinc sulfate. Test-positive samples were subjected to nested PCR amplification of the 18SSU ribosomal RNA gene for identification of Cryptosporidium and Giardia and of the gp60 gene for subtyping of Cryptosporidium. Microbiological analysis of water was carried out by the multiple-tube method and by means of a chromogenic substrate, and parasitological analysis was performed on 31 samples by direct immunofluorescence and nested PCR of the genes mentioned above. Identification of the species of Cryptosporidium was performed by sequencing and PCR with analysis of restriction fragment length polymorphisms. The prevalence of Giardia and Cryptosporidium was higher in calves than in adults. Among the samples of cattle feces, Cryptosporidium parvum was identified in 41 (64%, C. ryanae in eight (12.5%, C. bovis in four (6.3%, C. andersoni in five (7.8%, and a mixed infection in 20 samples (31.3%. These parasites were not identified in the samples of human feces. Thermotolerant coliform bacteria were identified in 25 samples of water (45.5%. Giardia duodenalis and C. parvum were identified in three water samples. The gp60 gene analysis of C. parvum isolates revealed the presence of two strains (IIaA20G1R1 and IIaA17G2R2 in the fecal samples and one (IIaA17G2R1 in the water samples. The presence of coliforms was associated with the water source, structure and degradation of springs, rain, and turbidity. The prevalence of protozoa was higher in calves up to six months of age. C. parvum and G
Toledo, Roberta Dos Santos; Martins, Felippe Danyel Cardoso; Ferreira, Fernanda Pinto; de Almeida, Jonatas Campos; Ogawa, Liza; Dos Santos, Hannah Lia Ettiene Peruch Lemos; Dos Santos, Maíra Moreira; Pinheiro, Filipe Aguera; Navarro, Italmar Teodorico; Garcia, João Luis; Freire, Roberta Lemos
The aims of this study were to verify the prevalence of Cryptosporidium spp. and Giardia spp. in animal feces and drinking water on dairy farms and to identify a possible relation between the exposure factors and the presence of these parasites. Fecal samples from cattle and humans and water samples were collected on dairy farms in Paraná, Brazil. Analysis of (oo)cysts in the feces was performed by the modified Ziehl-Neelsen staining and centrifugal flotation in zinc sulfate. Test-positive samples were subjected to nested PCR amplification of the 18SSU ribosomal RNA gene for identification of Cryptosporidium and Giardia and of the gp60 gene for subtyping of Cryptosporidium. Microbiological analysis of water was carried out by the multiple-tube method and by means of a chromogenic substrate, and parasitological analysis was performed on 31 samples by direct immunofluorescence and nested PCR of the genes mentioned above. Identification of the species of Cryptosporidium was performed by sequencing and PCR with analysis of restriction fragment length polymorphisms. The prevalence of Giardia and Cryptosporidium was higher in calves than in adults. Among the samples of cattle feces, Cryptosporidium parvum was identified in 41 (64%), C. ryanae in eight (12.5%), C. bovis in four (6.3%), C. andersoni in five (7.8%), and a mixed infection in 20 samples (31.3%). These parasites were not identified in the samples of human feces. Thermotolerant coliform bacteria were identified in 25 samples of water (45.5%). Giardia duodenalis and C. parvum were identified in three water samples. The gp60 gene analysis of C. parvum isolates revealed the presence of two strains (IIaA20G1R1 and IIaA17G2R2) in the fecal samples and one (IIaA17G2R1) in the water samples. The presence of coliforms was associated with the water source, structure and degradation of springs, rain, and turbidity. The prevalence of protozoa was higher in calves up to six months of age. C. parvum and G. duodenalis were
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
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
Guerrant. 2007. Heavy cryptosporidial infections in children in northeast Brazil: comparison of Cryptosporidium hominis and Cryptosporidium parvum...Asgharpour, C. T. Ng, D. P. Calfee, R. L. Guerrant, V. Maro, S. Ole-Nguyaine, and J. F. Shao. 2005. Short report: asymptomatic Cryptosporidium hominis ...JAN 2008 2. REPORT TYPE N/A 3. DATES COVERED - 4. TITLE AND SUBTITLE Intestinal immune response to human Cryptosporidium sp. infection 5a
Gómez-Couso, H; Fontán-Sainz, M; Fernández-Ibáñez, P; Ares-Mazás, E
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.
Robinson, Guy; Elwin, Kristin; Chalmers, Rachel M.
Several Cryptosporidium spp. are known to infect humans, but most cases of illness are caused by Cryptosporidium hominis or C. parvum. During a long-term genotyping in the United Kingdom, we identified 3 unusual Cryptosporidium genotypes (skunk, horse, and rabbit) in human patients with diarrhea.
Freppel, Wesley; Puech, Pierre-Henri; Ferguson, David J P; Azas, Nadine; Dubey, Jitender P; Dumètre, Aurélien
Toxoplasma gondii is a common parasite of humans and animals, which is transmitted via oocysts in cat faeces or tissue cysts in contaminated meat. The robust oocyst and sporocyst walls protect the infective sporozoites from deleterious external attacks including disinfectants. Upon oocyst acquisition, these walls lose their integrity to let the sporozoites excyst and invade host cells following a process that remains poorly understood. Given the resistance of the oocyst wall to digestive enzymes and the ability of oocysts to cause parenteral infections, the present study investigated the possible contribution of macrophages in supporting sporozoite excystation following oocyst internalisation. By using single cell micromanipulations, real-time and time-point imaging techniques, we demonstrated that RAW macrophages could interact rapidly with oocysts and engulfed them by remodelling of their actin cytoskeleton. Internalised oocysts were associated to macrophage acidic compartments and showed evidences of wall disruption. Sporozoites were observed in macrophages containing oocyst remnants or in new macrophages, giving rise to dividing tachyzoites. All together, these results highlight an unexpected role of phagocytic cells in processing T. gondii oocysts, in line with non-classical routes of infection, and open new perspectives to identify chemical factors that lead to oocyst wall disruption under physiological conditions.
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...
Anti-Cryptosporidium IgA and IgG are useful markers of exposure to Cryptosporidium in human populations, but detection in saliva may be difficult. To evaluate a magnetic microsphere assay for detection of anti-Cryptosporidium IgA and IgG in saliva, recombinant sporozoite gp15 (1...
Razakandrainibe, Romy; Diawara, El Hadji Ibrahima; Costa, Damien; Le Goff, Laetitia; Lemeteil, Denis; Ballet, Jean Jacques; Gargala, Gilles; Favennec, Loïc
Cryptosporidium spp. are infections the most frequent parasitic cause of diarrhea in humans and cattle. However, asymptomatic cases are less often documented than symptomatic cases or cases with experimentally infected animals. Cryptosporidium (C.) hominis infection accounts for the majority of pediatric cases in several countries, while C. parvum is a major cause of diarrhea in neonatal calves. In cattle Cryptosporidium spp. infection can be caused by C. parvum, C. bovis, C.andersoni and C. ryanae, and recently, reports of cattle cases of C. hominis cryptosporidiosis cases suggest that the presence of C. hominis in calves was previously underestimated. From February to November 2015, Cryptosporidium spp. infected calves were detected in 29/44 randomly included farms from 5 geographic regions of France. C. hominis and C. parvum were found in 12/44 and 26/44 farms, respectively with higher C. hominis prevalence in the western region. In 9 farms, both C. parvum and C. hominis were detected. Eighty-six of 412 (73/342 asymptomatic and 13/70 symptomatic) one to nine-week-old calves shed C. hominis or C. parvum oocysts (15 and 71 calves, respectively), with no mixed infection detected. The predominant C. hominis IbA9G3 genotype was present in all regions, and more frequent in the western region. An incompletely characterized Ib, and the IbA13G3, IbA9G2 and IbA14G2 genotypes were present only in the western region. For C. parvum, the most frequent genotype was IIaA16G3R1 with no geographic clustering. Most C. hominis infected calves were asymptomatic, with some exceptions of IbA9G2 and IbA9G3 isolates, while C. parvum IIaA16G3R1 was associated with symptoms. Present results indicate for the first time that in several geographic regions of France, C. hominis was present in about one fifth of both asymptomatic and symptomatic infected calves, with isolated genotypes likely associated with human infection. Further investigations are aimed at documenting direct or indirect
Ruiz, Antonio; Muñoz, María Carmen; Molina, José Manuel; Hermosilla, Carlos; Andrada, Marisa; Lara, Pedro; Bordón, Elisa; Pérez, Davinia; López, Adassa María; Matos, Lorena; Guedes, Aránzazu Carmen; Falcón, Soraya; Falcón, Yaiza; Martín, Sergio; Taubert, Anja
Caprine coccidiosis, affecting mainly young goat kids around the weaning period, is worldwide the most important disease in the goat industry. Control of caprine coccidiosis is increasingly hampered by resistances developed against coccidiostatic drugs leading to an enhanced need for anticoccidial vaccines. In the current study we conducted an oral immunization trial with live attenuated sporulated Eimeria ninakohlyakimovae oocysts. Sporulated E. ninakohlyakimovae oocysts were attenuated by X-irradiation technique. The experimental design included a total of 18 goat kids divided into the following groups: (i) animals immunized with attenuated E. ninakohlyakimovae oocysts at 5 weeks of age and challenged 3 weeks later with non-irradiated homologous oocysts (group 1); (ii) animals infected with non-attenuated E. ninakohlyakimovae oocysts at 5 weeks of age and challenged 3 weeks later with non-attenuated homologous oocysts (group 2); (iii) animals primary-infected with untreated E. ninakohlyakimovae oocysts at 8 weeks of age (control of the challenge infection, group 3); (iv) non-infected control animals (group 4). Goat kids immunized with live attenuated E. ninakohlyakimovae oocysts (group 1) excreted significantly less oocysts in the faeces (95.3% reduction) than kids infected with non-attenuated ones (group 2). Furthermore, immunization with live but attenuated oocysts resulted in ameliorated clinical coccidiosis compared to goat kids infected with untreated oocysts (group 2) and resulted in equally reduced signs of coccidiosis after challenge infection compared to acquired immunity driven by non-attenuated oocysts. Overall, the present study demonstrates for the first time that live attenuated E. ninakohlyakimovae oocysts orally administered showed almost no pathogenicity but enough immunogenicity in terms of immunoprotection. Importantly, vaccinated animals still shed low amounts of oocysts, guaranteeing environmental contamination and consecutive booster
Full Text Available Introduction & Objective: Cryptosporidiosis caused by Cryptosporidium, which is a protozoan parasite, has a worldwide distribution. The infection is through fecal-oral route, direct or indi-rect contact, food or water. The treatment of cryptosporidiosis is difficult and the anti-parasitic agents are not effective. The purpose of this study was encapsulation of nitazoxanide in solid lipid nano-particles (SLN and investigation of its anti-Cryptosporidium effect and its comparison with free drug in the neonatal rat. Materials & Methods: Nitazoxanide was encapsulated by HPH method with 2 mg/Kg concentra-tion in SLN nanoparticles. The oocysts were collected from calves and purified by sucrose floatation. A total of 72 Wistar neonatal rats were categorized in 6 groups of 12 rats including four infected groups treated by free drug, encapsulated nano drug, colloidal carriers without drug (SLN and olive oil; an infected control group and a healthy control group that received PBS. 5 × 105 of oocyts inoculated orally into the sample groups. Finally, intestine of each rat was homogenized in PBS by rotor and the homogenized material was passed through a sieve. Then, floated oocysts in sucrose solution were counted by hemocytometer. Results: Treatment by nitazoxanide significantly decreased the number of parasites in the treatment groups. This decrease at day 6 was more than day 3. Nano nitazoxanide had more effects on parasites than free drug. This difference at day 3 of treatment was not significant (p= 0.182 but at day 6 was statistically significant (P< 0.001. Conclusion: Using nano-nitazoxanide could be a more effective way in the treatment of Cryp-tosporidium infections. (Sci J Hamadan Univ Med Sci 2016; 23 (2:134-140
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...
Alonso Molina, José Luís; Amoros-Muñoz, Inmaculada; Guy, Rebecca A.
Real-time PCR (qPCR) is a rapid tool to quantify pathogens in the aquatic environment; however, it quantifies all pathogens, including both viable and nonviable. Propidium monoazide (PMA) is a membrane-impairment dye that penetrates only membrane-damaged cells. Once inside the cell, PMA is covalently cross-linked to DNA through light photoactivation, and PCR amplification is strongly inhibited. The goal of this study was to evaluate PMA-qPCR assays for rapid quantifica...
Robertson, L J; Gjerde, B
While recently published techniques for recovering parasites from fruits and vegetables demonstrate a marked increase in efficiency and utility, there is still scope for further improvement in developing a standard method, particularly with difficult, but important, sample matrices such as bean sprouts. Herein, a number of parameters used in published techniques are investigated more closely. While sample size reduction may improve recovery efficiency because of a range of factors, it is important to keep the sample large enough for detection of low-level contamination. Age of sample is also important, and samples should be as fresh as possible. Elution procedures may contribute to losses of Giardia and should be more thoroughly investigated. Improved immunomagnetic separation techniques currently coming onto the market also have the potential to increase recovery efficiency substantially, even with difficult samples such as aged bean sprouts. However, merely increasing magnetic strength of the capturing magnet does not affect recovery efficiency, which must be reliant on a superior bead system, buffering system, or both.
The purpose of this study was to determine if Eimeria oocyst concentrations and species composition in commercial broiler house litter changed during different cycles of anticoccidial drug (ACD) or live Eimeria oocyst vaccine (VAC) control programs, and if there was a correlation between Eimeria ooc...
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.
Gómez-Couso, H; Fontán-Sainz, M; Navntoft, C; Fernández-Ibáñez, P; Ares-Mazás, E
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. Copyright © 2012 Royal Society of Tropical Medicine and Hygiene. Published by Elsevier Ltd. All rights reserved.
Certad, Gabriela; Viscogliosi, Eric; Chabé, Magali; Cacciò, Simone M
Intestinal protozoa are important etiological agents of diarrhea, particularly in children, yet the public health risk they pose is often neglected. Results from the Global Enteric Multicenter Study (GEMS) showed that Cryptosporidium is among the leading causes of moderate to severe diarrhea in children under 2 years. Likewise, Giardia infects approximately 200 million individuals worldwide, and causes acute diarrhea in children under 5 years. Despite this recognized role as pathogens, the question is why and how these parasites cause disease in some individuals but not in others. This review focuses on known pathogenic mechanisms of Cryptosporidium and Giardia, and infection progress towards disease. Copyright © 2017 Elsevier Ltd. All rights reserved.
Full Text Available Aim: The objective of this study was to investigate prevalence of gastrointestinal parasites and Cryptosporidium species in extensively managed pigs in Mekelle and urban areas of southern zone of Tigray Region, Ethiopia during June - September, 2012. Material and methods: Seven hundred fourteen pigs of different ages and sexes were selected for fecal sample collection. Fecal samples were collected from the rectum of pigs with strict sanitation. A total of 25 soil samples were also collected from backyards of pig pens using clean zipped plastic bags. Both fecal and soil samples were examined for eggs and cysts of GIT parasites by flotation and sedimentation techniques. Modified Ziehl Neelsen technique was used to examine oocysts of Cryptosporidium species from 276 randomly selected fecal samples. Results: Out of 714 pigs examined through flotation and sedimentation, 27.3% were infected by at least one gastrointestinal parasite. Ascaris suum (25.9% was the most prevalent parasite followed by Fasciola hepatica (1.8%, Eimeria spp. (1.7% and Trichuris suis (0.3%. There was no significant association between sex and prevalence of parasites ÷2[df 1] = 1.921; P=0.166. Contrary to this, age of pigs had effect on prevalence of parasites ÷2[df 2] = 8.376; P=0.015. About 7% of pigs examined were positive for oocysts of Cryptosporidium spp. Moreover, 72% of the soil samples found to be contaminated with eggs of Ascaris spp. in the study area. Apart from causing morbidity in the infected pigs, the potential of Ascaris of pigs to infect man and vice versa together with poor environmental hygiene, may complicate the epidemiology and control of Ascariasis in the study areas. Extensively managed pigs may also act as potential reservoirs for zoonoses of Cryptosporidium species. Conclusion: It is concluded that further investigations are crucial on molecular characterization of Ascaris and Cryptosporidium isolates of extensively managed pigs to determine the
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
Metge, D.W.; Harvey, R.W.; Aiken, G.R.; Anders, R.; Lincoln, G.; Jasperse, James; Hill, M.C.
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. parvumoocysts and oocyst-sized (3 μm) carboxylate-modified microspheres through poorly sorted (sorting indices, σ1, 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 judging from high α 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.
Evaluation and Comparison of Enzyme Immunoassay (Eia and Acid Fast Staining with Confirmation by Immunofluorescent Antibody Assay for Detection of Cryptosporidium Species in Infants and Young Children.
D Dorostcar Moghaddam
Full Text Available Introduction: Cryptosporidiosis is prevalent world wide, causing a variety of problems ranging from acute, self-limiting diarrhea to fatal cases in immunocompromised persons, particulary those with acquired immunodeficiency (AIDS. Diagnosis of Cryptosporidium is made by identification of oocysts in stool specimens. The detection is most commonly made by the acid-fast staining method followed by microscopic examination which has low specificity and sensitivity. Material and Methods: In the present study, we evaluated diagnostic utility of a commercially available enzyme immunoassay (EIA, which detects Cryptosporidium-Specific antigen (CSA in 204 unprocessed stool specimens obtained from patients less than 3 years of age. Results: When compared with the routine screening procedure applied in this field study (screening by acid-fast staining and microscopy after concentration of positive results by IFA, both sensitivity and specificity were 98%. Of the 139 specimens negative by microscopy, 13 (9.3% were positive by EIA, 11 of which were confirmed by inhibition with antibody to Cryptosporidia-specific antigen. Conclusion: The EIA is an important tool for identifying Cryptosporidium in fecal specimens in field studies since it is sensitive, specific, simple to use and unaffected by the presence of a preservative.
Siwila, J.; Phiri, I.G.K.; Enemark, Heidi L.
Prevalence, incidence and seasonal variation of Cryptosporidium and Giardia duodenalis were studied over a 12-month period in 100 children from four pre-schools in Kafue, Zambia. Questionnaire data and a single stool sample were collected monthly from each child. Samples were processed using...... a commercial kit (Meridian Diagnostics Inc., USA) and oo(cysts) visualised by immunofluorescence microscopy. Cryptosporidium was detected in 30.7% (241/786; 95% CI = 27.5-33.9) while G. duodenalis was detected in 29.0% (228/786; 95% CI = 25.8-32.2). A total of 86% experienced one or more episodes...... of cryptosporidiosis while 75% had giardiasis. Cumulative incidence per 100 children was 75.4 for Cryptosporidium and 49.0 for G. duodenalis. Both infections were significantly more common in the wet compared to the dry season (34.8%, 162/466 vs. 24.7%, 79/320, P = 0.003 and 35.2%, 164/466 vs. 20.0%, 64/320, P
Kelley, M.B. [United States Military Academy, West Point, NY (United States); Brokaw, J.K. [Rensselaer Polytechnic Institute, Troy, NY (United States); Brokaw, J.K. [Army Center for Health Promotion and Preventive Medicine, Aberdeen Proving Ground, MD (United States); Warrier, P.K. [Rensselaer Polytechnic Institute, Troy, NY (United States)] [and others
This paper provides the results of a study of two U.S. Army installation drinking water sources and treatment systems for the removal of Giardia and Cryptosporidium. Sampling was conducted monthly for one year commencing in late March 1994 and concluding at the end of February 1995. Results of this detailed study include examination of turbidity, particle counts, and total and fecal coliforms as well as the enumeration of Giardia cysts and Cryptosporidium oocysts. Our goal, in addition to determining the levels of these pathogens in both raw and product waters, was to determine if typical operating parameters would be helpful in identifying either elevated raw water protozoa or breakthrough of either pathogen in the product water from the treatment facilities. A data summary for the results of the protozoa enumeration is a Table 1. Our results indicate frequent contamination of the raw waters at both sites by either or both pathogens. Further, we observed sporadic breakthrough of low levels of Cryptosporidium in the filtered waters of both sites. The method employed to concentrate, purify and enumerate the pathogenic protozoa is also discussed and comparisons are made to the proposed Information Collection Rule (ICR) method for detection of these microorganisms.
Segura, Remedios; Prim, Núria; Montemayor, Michel; Valls, María Eugenia; Muñoz, Carme
Cryptosporidium infection is a worldwide cause of diarrheal disease. To gain insight into the epidemiology of the infection in a certain geographic area, molecular methods are needed to determine the species/genotypes and subtypes. From 2004 to 2009, 161 cryptosporidiosis cases were detected in two hospitals in Barcelona. Diagnosis was performed by microscopic observation of oocysts in stool specimens following modified Ziehl-Neelsen staining. Most cases (82%) occurred in children. The number of cases increased in summer and autumn. Molecular characterization of Cryptosporidium was performed in 69 specimens, and C. hominis and C. parvum were identified in 88.4% and 10.1% of the cases, respectively. C. meleagridis was detected in one specimen. Subtyping based on the gp60 polymorphism showed six subtypes, four C. hominis and two C. parvum. Subtype IbA10G2 was the most prevalent subtype corresponding to 90% of all C. hominis isolates. This is the first report on the distribution of specific Cryptosporidium subtypes from humans in Spain. In our geographic area, the anthroponotic behavior of C. hominis, the lower infective dose, and the higher virulence of certain subtypes may contribute to the high incidence of human cryptosporidiosis caused by the IbA10G2 subtype. Further studies should include populations with asymptomatic shedding of the parasite.
Full Text Available Cryptosporidium infection is a worldwide cause of diarrheal disease. To gain insight into the epidemiology of the infection in a certain geographic area, molecular methods are needed to determine the species/genotypes and subtypes.From 2004 to 2009, 161 cryptosporidiosis cases were detected in two hospitals in Barcelona. Diagnosis was performed by microscopic observation of oocysts in stool specimens following modified Ziehl-Neelsen staining. Most cases (82% occurred in children. The number of cases increased in summer and autumn. Molecular characterization of Cryptosporidium was performed in 69 specimens, and C. hominis and C. parvum were identified in 88.4% and 10.1% of the cases, respectively. C. meleagridis was detected in one specimen. Subtyping based on the gp60 polymorphism showed six subtypes, four C. hominis and two C. parvum. Subtype IbA10G2 was the most prevalent subtype corresponding to 90% of all C. hominis isolates. This is the first report on the distribution of specific Cryptosporidium subtypes from humans in Spain.In our geographic area, the anthroponotic behavior of C. hominis, the lower infective dose, and the higher virulence of certain subtypes may contribute to the high incidence of human cryptosporidiosis caused by the IbA10G2 subtype. Further studies should include populations with asymptomatic shedding of the parasite.
Němejc, K.; Sak, Bohumil; Květoňová, Dana; Hanzal, V.; Janiszewski, P.; Forejtek, P.; Rajský, D.; Ravaszová, P.; McEvoy, J.; Kváč, Martin
Roč. 197, 3-4 (2013), s. 504-508 ISSN 0304-4017 Grant - others:Jihočeská univerzita(CZ) 022/2010/Z; Jihočeská univerzita(CZ) 11/2013/Z Institutional support: RVO:60077344 Keywords : Central Europe * Cryptosporidium scrofarum * Cryptosporidium suis * Eurasian wild boar * PCR * SSU Subject RIV: EE - Microbiology, Virology Impact factor: 2.545, year: 2013
Hashim, Amna; Mulcahy, Grace; Bourke, Billy; Clyne, Marguerite
Cryptosporidiosis in humans is caused by the zoonotic pathogen Cryptosporidium parvum and the anthroponotic pathogen Cryptosporidium hominis. To what extent the recently recognized C. hominis species differs from C. parvum is unknown. In this study we compared the mechanisms of C. parvum and C. hominis invasion using a primary cell model of infection. Cultured primary bovine and human epithelial intestinal cells were infected with C. parvum or C. hominis. The effects of the carbohydrate lecti...
Sheoran, Abhineet; Wiffin, Anthony; Widmer, Giovanni; Singh, Pradeep; Tzipori, Saul
Cryptosporidium hominis and Cryptosporidium parvum, which infect humans equally, are genetically/antigenically almost identical. It remains unclear, however, whether infection with C. hominis protects against C. parvum. Gnotobiotic piglets were used to investigate cross-protection. After ≥3 days of recovery from C. hominis infection, the piglets were completely protected against subsequent challenge with C. hominis but only partially against challenge with C. parvum, as compared with age-m...
Maddox-Hyttel, Charlotte; Langkjær, Rikke Breinhold; Enemark, Heidi L.
and sow herds, respectively. Each herd was visited once for the collection of faecal samples and registration of basic management parameters. Faecal samples were collected from three different age groups of animals, i.e. 5 sows/cows. 10 nursing piglets/calves less than 1 month, and 10 weaner pigs 8-45 kg....../calves 1-12 months. The faecal samples were purified and the number of (oo)cysts quantified. The study revealed an age-specific herd prevalence of Cryptosporidium of 16, 31 and 100% for sows, piglets and weaners, respectively, and of 14, 96 and 84% for cows, young calves and older calves, respectively....... For Giardia the age-specific herd prevalence was 18, 22 and 84% for the sows, piglets and weaners, while for cattle herds the prevalence was 60, 82 and 100% for cows, young calves and older calves, correspondingly. The (oo)cyst excretion levels varied considerably both within and between herds for all age...
Petterson, S; Roser, D; Deere, D
It is proposed that the next revision of the Australian Drinking Water Guidelines will include 'health-based targets', where the required level of potable water treatment quantitatively relates to the magnitude of source water pathogen concentrations. To quantify likely Cryptosporidium concentrations in southern Australian surface source waters, the databases for 25 metropolitan water supplies with good historical records, representing a range of catchment sizes, land use and climatic regions were mined. The distributions and uncertainty intervals for Cryptosporidium concentrations were characterized for each site. Then, treatment targets were quantified applying the framework recommended in the World Health Organization Guidelines for Drinking-Water Quality 2011. Based on total oocyst concentrations, and not factoring in genotype or physiological state information as it relates to infectivity for humans, the best estimates of the required level of treatment, expressed as log10 reduction values, ranged among the study sites from 1.4 to 6.1 log10. Challenges associated with relying on historical monitoring data for defining drinking water treatment requirements were identified. In addition, the importance of quantitative microbial risk assessment input assumptions on the quantified treatment targets was investigated, highlighting the need for selection of locally appropriate values.
Kváč, Martin; Květoňová, Dana; Sak, Bohumil; Ditrich, Oleg
Roč. 15, č. 6 (2009), s. 982-983 ISSN 1080-6040 R&D Projects: GA ČR GP523/07/P117 Institutional research plan: CEZ:AV0Z60220518 Keywords : immunocompetent patients * cryptosporidiosis * Cryptosporidium pig genotype II Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 6.794, year: 2009
Objective: The objective of this study is to search for Cryptosporidium parvum in Sudanese immunocompromised and immunocompetent patients presenting with diarrhea. Methods: Two hundred and thirteen stool specimens were collected from different groups of patients presenting with diarrhea and healthy control ...
Schrijven JF; Bruin HAM de; Engels GB; Leenen EJTM; MGB
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
Zhao, Wei; Wang, Rongjun; Zhang, Weizhe; Liu, Aiqin; Cao, Jianping; Shen, Yujuan; Yang, Fengkun; Zhang, Longxian
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.
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.
Full Text Available 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.
Liu, Xuehan; Xie, Na; Li, Wei; Zhou, Ziyao; Zhong, Zhijun; Shen, Liuhong; Cao, Suizhong; Yu, Xingming; Hu, Yanchuan; Chen, Weigang; Peng, Gangneng
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.
Full Text Available Cryptosporidium is one of the most important parasites in poultry, and this pathogen can infect more than 30 avian species. The present study investigated the infection rate of Cryptosporidium among broiler chicken flocks. A total of 385 fecal samples from broiler chickens in 7 regions of Zhejiang Province collected from November 2010 to January 2012 were examined by microscopy. Thirty-eight (10% samples were positive for Cryptosporidium infection, and 3 genotypes (Cryptosporidium baileyi, Cryptosporidium meleagridis, and avian genotype II were identified by PCR and sequencing. A phylogenetic tree of the isolates was analyzed. These results suggest that cryptosporidiosis is widespread in poultry in Zhejiang Province, and is a potential threat to public health as well as the economy. This is the first report about the infection rate and molecular characterization of Cryptosporidium in broiler chickens in Zhejiang.
Full Text Available Plasmodium sporozoites develop within oocysts residing in the mosquito midgut. Mature sporozoites exit the oocysts, enter the hemolymph, and invade the salivary glands. The circumsporozoite (CS protein is the major surface protein of salivary gland and oocyst sporozoites. It is also found on the oocyst plasma membrane and on the inner surface of the oocyst capsule. CS protein contains a conserved motif of positively charged amino acids: region II-plus, which has been implicated in the initial stages of sporozoite invasion of hepatocytes. We investigated the function of region II-plus by generating mutant parasites in which the region had been substituted with alanines. Mutant parasites produced normal numbers of sporozoites in the oocysts, but the sporozoites were unable to exit the oocysts. In in vitro as well, there was a profound delay, upon trypsin treatment, in the release of mutant sporozoites from oocysts. We conclude that the exit of sporozoites from oocysts is an active process that involves the region II-plus of CS protein. In addition, the mutant sporozoites were not infective to young rats. These findings provide a new target for developing reagents that interfere with the transmission of malaria.
Abouel-Nour, Mohamed F; El-Shewehy, Dina Magdy M; Hamada, Shadia F; Morsy, Tosson A
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.
U.S. Environmental Protection Agency — The data set is a spreadsheet that contains results of inactivation experiments that were conducted to to determine the effectiveness of chlorine in inactivating B....
Wang, Andrea; Ruch-Gallie, Rebecca; Scorza, Valeria; Lin, Philip; Lappin, Michael R
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. Copyright © 2011 Elsevier B.V. All rights reserved.
Al-Mathal, Ebtisam M; Alsalem, Afaf M
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 (Pgranatum-treated mice was significantly higher than that of the infected/untreated controls throughout the study (Pgranatum-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
Kinsey, Erin; Korte, Caroline; L'Ollivier, Coralie; Dubey, Jitender; Dumetre, Aurélien; Darnault, Christophe
Toxoplasma gondii has a complex lifecycle that involves a wide variety of intermediate hosts with felids as the definitive host. Because of its numerous hosts and the prevalence of cats, T.gondii has spread throughout nearly the entire globe. Oocysts have been found not only in the feces of cats, but also in soils, animal feeds and water. Exposure through consumption of infected meat or following contact with cat feces can cause damage to the eyes, brain and other organs of immunocompromised populations as well as fetuses if they are exposed in utero. The prevalence of T.gondii and potential health risks necessitate a better understanding of the transport of T.gondii through soils, which to this point has not been well studied. This work aims to characterize the transport and retention of T.gondii oocysts in a number of unsaturated natural soils where fast transport and preferential flow paths have been prevented. The soils used are classified as loamy sands and sandy loams. They were placed in soil columns at a known bulk density and were then subjected to an artificial rain of 1 mM KCl solution. Flow in the columns was vertical and gravity driven. After steady state was reached, a pulse containing 2.5 million T.gondii oocysts and KBr as a conservative tracer was applied to the top of the column, after which steady rainfall was resumed. Leachate samples were collected throughout the experiment. qPCR for T.gondii was performed and KBr ions were measured to create breakthrough curves for both. After the completion of the rainfall portion of the experiment, soil columns were cut into 1 to 2 cm sections and analyzed for T.gondii with qPCR to characterize retention within the column and for soil water content.
Mirhashemi, Marzieh Ezzaty; Zintl, Annetta; Grant, Tim; Lucy, Frances; Mulcahy, Grace; De Waal, Theo
Cryptosporidium is a protozoan that can cause gastro-intestinal illness with diarrhoea in a wide range of hosts. In fact some species of Cryptosporidium can infect the broad range of hosts. The current paper is focused to investigate monthly prevalence and diversity of Cryptosporidium spp. during the spring and early summer (March-June) in 2009 and 2010 in farms with no history of cryptosporidiosis. Animal samples were analyzed to elucidate the prevalence of Cryptosporidium in two regions, West and the East catchments in Ireland. Our investigation demonstrates the prevalence ranges from 14% to 26% an early summer peak (June) was observed. Based on the findings of this study Cryptosporidium ryanae (in cattle, horses), and Cryptosporidium bovis/xiaoi followed by Cryptosporidium parvum (in sheep) were found to be the predominant species in asymptomatic cases. The circulation of other Cryptosporidium species such as C. parvum, C. bovis, C. ubiquitum, C. andersoni and Cryptosporidium horse and pig genotypes in livestock was investigated. Published by Elsevier B.V.
Prediger, Jitka; Horčičková, Michaela; Hofmannová, Lada; Sak, Bohumil; Ferrari, Nicola; Mazzamuto, Maria Vittoria; Romeo, Claudia; Wauters, Lucas A; McEvoy, John; Kváč, Martin
The present study was undertaken to describe Cryptosporidium spp. infection in tree squirrels from 17 locations in Northern Italy. A total of 357 squirrels were examined, including species native to Europe (Sciurus vulgaris; n=123), and species introduced from North America (Sciurus carolinensis; n=162) and Southeast Asia (Callosciurus erythraeus; n=72). Faecal samples of all squirrels were examined for the presence of Cryptosporidium infection by microscopy (flotation method) and PCR/sequence analysis of the Cryptosporidium 18S rRNA, actin, and gp60 genes. Despite the overlapping ranges of native and introduced tree squirrel species in the study area, they host different Cryptosporidium spp. Sciurus vulgaris were exclusively infected with Cryptosporidium ferret genotype (n=13) belonging to three novel gp60 subtypes, VIIIb-VIIId. Sciurus carolinensis hosted C. ubiquitum subtype XIIb (n=2), Cryptosporidium skunk genotype subtype XVIa (n=3), and chipmunk genotype I subtype XIVa (n=1). Cryptosporidium chipmunk genotype I subtype XIVa was also found in two C. erythraeus. Comparing data from this and previous studies, we propose that Cryptosporidium skunk genotype and possibly C. ubiquitum subtype XIIb were introduced to Europe with eastern grey squirrels. Cryptosporidium chipmunk genotype I and ferret genotype were associated with high intensity infections, but there was no association with diarrhoea. Copyright © 2017 Elsevier GmbH. All rights reserved.
Fetterer, Raymond H; Jenkins, Mark C; Miska, Katarzyna B; Barfield, Ruth C
The current study investigates the use of irradiated oocysts to protect broiler chicks, raised on litter, from infection with multiple species of Eimeria. In order to determine the optimum radiation dose for each Eimeria species, 1-day-old chicks were immunized with oocysts of Eimeria maxima, Eimeria acervulina, or Eimeria tenella exposed to gamma radiation ranging from 0-500 Gy. The litter oocyst counts at 7 days postimmunization, and the effect on weight gain following a challenge infection, decreased with an optimum dose between 150-200 Gy. Based on this finding, broiler chicks were immunized with a mixture of E. maxima, E. acervulina, and E tenella that had been exposed to 150 or 200 Gy. This resulted in more than a 100-fold reduction in litter oocyst counts and significant protection from a challenge infection, as measured by improved weight gain and feed conversion ratio (FCR). Immunization of birds with oocysts receiving 200 Gy was less effective in providing protection from a challenge infection. An additional formulation of vaccines containing two different oocyst doses of the three species that had been irradiated with 150 Gy were evaluated in their ability to attenuate oocyst output and convey protection to challenge. Results were similar with both high and low numbers of irradiated oocysts. Immunized chicks shed less oocysts at 7 days postimmunization and were protected from negative effects of challenge infection as measured by FCR, changes in weight gain, lesion scores, and measurement of body composition. However, the level of protection was somewhat less than that achieved by immunization with nonirradiated oocysts. The overall conclusion is that an irradiated oocyst vaccine developed in this study can effectively protect chicks that are raised on litter from challenge infection with multiple species of Eimeria, comparable to vaccines with virulent or precocious strains.
Toxoplasma gondii is a common parasite of humans and domestic animals, which is transmitted via oocysts in cat faeces or tissue cysts in contaminated meat. The oocyst and sporocyst walls are multilayered polymeric structures that protect the infective sporozoites from deleterious physical and chemic...
Full Text Available The Caliciviridae family of viruses contains clinically important human and animal pathogens, as well as vesivirus 2117, a known contaminant of biopharmaceutical manufacturing processes employing Chinese hamster cells. An extensive literature exists for inactivation of various animal caliciviruses, especially feline calicivirus and murine norovirus. The caliciviruses are susceptible to wet heat inactivation at temperatures in excess of 60 °C with contact times of 30 min or greater, to UV-C inactivation at fluence ≥30 mJ/cm2, to high pressure processing >200 MPa for >5 min at 4 °C, and to certain photodynamic inactivation approaches. The enteric caliciviruses (e.g.; noroviruses display resistance to inactivation by low pH, while the non-enteric species (e.g.; feline calicivirus are much more susceptible. The caliciviruses are inactivated by a variety of chemicals, including alcohols, oxidizing agents, aldehydes, and β-propiolactone. As with inactivation of viruses in general, inactivation of caliciviruses by the various approaches may be matrix-, temperature-, and/or contact time-dependent. The susceptibilities of the caliciviruses to the various physical and chemical inactivation approaches are generally similar to those displayed by other small, non-enveloped viruses, with the exception that the parvoviruses and circoviruses may require higher temperatures for inactivation, while these families appear to be more susceptible to UV-C inactivation than are the caliciviruses.
Catalina Avendaño V
Full Text Available Cryptosporidium is a zoonotic parasite very important in animal health as well as in public health. It is because this is one of the main causes of diarrhea in children, calves, lambs and other variety of youth mammalians in a lot of countries. The globalization has enabled the exchange of biological material in different regions worldwide, encouraging the spread of diseases and exposure to these biological agents to different environmental conditions, inducing adaptation through genetic changes. Based in the polymorphism of the gene for GP60, this review intended to present the distribution of Cryptosporidium parvum and Cryptosporidium hominis in humans and calves worldwide. The subtype that affects cattle more frequently corresponds to IIaA15G2R; while the subtype most frequently isolated from human samples is IaA19G2.
Full Text Available Intrinsic vector characteristics and environmental factors affect the sporogonic development of P. falciparum in Anopheles mosquitoes. We tested for the presence of the circumsporozoite protein, as a marker of the oocyst to sporozoite transition in naturally infected Anopheles gambiae s.l. and Anopheles funestus. Malaria vectors were collected in a village in the Sahel of Niger during the rainy and dry seasons. ELISA-CSP was carried out on abdomen and head/thorax portions from more than 2,000 samples. No significant difference was found in the overall rates of infection of An. gambiae s.l. (4.13% and An. funestus (3.58%. Given the differences in duration of the two parasite stages, P. falciparum CSP antigen prevalence was nearly as high in the abdomen as in the head/thorax, and did not differ significantly between An. gambiae s.l. and An. funestus. These preliminary results suggest that development from oocysts to salivary gland sporozoites is similar in the two vectors. However, these developmental indices varied as a function of the season in which samples were collected, particularly for An. gambiae s.l. This simple method may be useful for field studies assessing the effect of environmental and genetic factors on parasite survival.
Full Text Available The objective of this study was to find out the impact of environmental conditions on the survival of intestinal parasites on environmental surfaces commonly implicated in the transmission of these parasites. The study was performed by incubating Cryptosporidium and Giardia (oocysts on environmentally relevant surfaces such as brushed stainless steel, formica, ceramic, fabric, and skin. Parallel experiments were conducted using clean and soiled coupons incubated under three temperatures. The die-off coefficient rates (K were calculated using first-order exponential formula. For both parasites, the fastest die-off was recorded on fabric, followed by ceramic, formica, skin, and steel. Die-off rates were directly correlated to the incubation temperatures and surface porosity. The presence of organic matter enhanced the survivability of the resting stages of test parasites. The decay rates calculated in this study can be used in models for public health decision-making process and highlights the mitigation role of hand hygiene agents in their prevention and control.
Full Text Available Cryptosporidium spp. are coccidians, oocysts-forming apicomplexan protozoa, which complete their life cycle both in humans and animals, through zoonotic and anthroponotic transmission, causing cryptosporidiosis. The global burden of this disease is still underascertained, due to a conundrum transmission modality, only partially unveiled, and on a plethora of detection systems still inadequate or only partially applied for worldwide surveillance. In children, cryptosporidiosis encumber is even less recorded and often misidentified due to physiological reasons such as early-age unpaired immunological response. Furthermore, malnutrition in underdeveloped countries or clinical underestimation of protozoan etiology in developed countries contribute to the underestimation of the worldwide burden. Principal key indicators of the parasite distribution were associated to environmental (e.g., geographic and temporal clusters, etc. and host determinants of the infection (e.g., age, immunological status, travels, community behaviours. The distribution was geographically mapped to provide an updated picture of the global parasite ecosystems. The present paper aims to provide, by a critical analysis of existing literature, a link between observational epidemiological records and new insights on public health, and diagnostic and clinical impact of cryptosporidiosis.
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.
Hadfield, Stephen J.; Jackson, Colin J.; Elwin, Kristin; Xiao, Lihua; Hunter, Paul
UK Cryptosporidium hominis isolates have previously shown slight PCR fragment length polymorphism at multiple loci. To further investigate transmission, we conducted a case–control study and sequenced the GP60 locus from 115 isolates. Nine subtypes were identified; IbA10G2 predominated. Having a non-IbA10G2 subtype was significantly linked to recent travel outside Europe. PMID:18325272
Sheoran, Abhineet; Wiffin, Anthony; Widmer, Giovanni; Singh, Pradeep; Tzipori, Saul
Cryptosporidium hominis and Cryptosporidium parvum, which infect humans equally, are genetically/antigenically almost identical. It remains unclear, however, whether infection with C. hominis protects against C. parvum. Gnotobiotic piglets were used to investigate cross-protection. After ≥3 days of recovery from C. hominis infection, the piglets were completely protected against subsequent challenge with C. hominis but only partially against challenge with C. parvum, as compared with age-matched control animals challenged with either species. In conclusion, C. hominis-specific immunity was sufficient to completely protect against challenge with the same species but insufficient to provide the same level of protection against C. parvum.
Jenkins, Mark C; Parker, Carolyn; Klopp, Spangler; O'Brien, Celia; Miska, Katarzyna; Fetterer, Raymond
Vaccines composed of either virulent or attenuated Eimeria spp. oocysts have been developed as an alternative to medication of feed with ionophore drugs or synthetic chemicals. The purpose of this study was to evaluate the use of gel-beads containing a mixture of Eimeria acervulina, Eimeria maxima, and Eimeria tenella oocysts as a vaccine against coccidiosis. Newly hatched chicks (Gallus gallus domesticus) were either sprayed with an aqueous suspension of Eimeria oocysts or were allowed to ingest feed containing Eimeria oocysts-incorporated gel-beads. Control day-old chicks were given an equivalent number of Eimeria oocysts (10(4) total) by oral gavage. After 3 days, chicks were randomly assigned to individual cages, and feces were collected between days 5 and 8 postinfection. All samples were processed for total Eimeria oocysts. At 4 wk of age, all chickens and a control nonimmunized group received a high-dose E acervulina, E maxima, and E. tenella challenge infection. Oocyst excretion by chicks fed gel-beads or inoculated by oral gavage was 10- to 100-fold greater than that of chicks spray-vaccinated with the Eimeria oocysts mixture (log 6.3-6.6 vs. log 4.8). Subsequent protection against challenge as measured by weight gain and feed conversion efficiency was significantly greater (P 0.05) in weight gain and feed conversion efficiency compared with nonchallenged controls. These findings indicate that incorporation of Eimeria spp. oocysts in gel-beads may represent an effective way to deliver live oocyst vaccines to day-old chicks for preventing subsequent outbreaks of coccidiosis in the field.
Despite the health risks associated with exposure to Cryptosporidium and Giardia, there is no uniform approach to monitoring these protozoan parasites across the world. In the present study, a strategy for monitoring Cryptosporidium and Giardia in drinking water was developed in an effort to ensure that the risk of exposure ...
Kváč, Martin; Hofmannová, L.; Hlásková, Lenka; Květoňová, Dana; Vitovec, J.; McEvoy, J.; Sak, Bohumil
Roč. 201, 1-2 (2014), s. 9-17 ISSN 0304-4017 R&D Projects: GA MŠk(CZ) LH11061 Institutional support: RVO:60077344 Keywords : Cryptosporidium erinacei * taxonomy * morphology * molecular analyses * transmission studies * Cryptosporidium hedgehog genotype Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 2.460, year: 2014
One hundred and eleven of 1229 children (9%) aged < 10 years admitted to King Edward VIII Hospital, Durban, with gastro-enteritis over a period of 1 year were found to harbour Cryptosporidium. Of these, 96 (89,7%) were < 2 years of age. Cryptosporidium was the only potential pathogen identified in 80 of these patients ...
The prevalence of Cryptosporidium spp. in cattle faeces in Ogun state, Nigeria was determined by a commercially produced enzyme-linked immunosorbent assay kit. Out of a total of 200 samples, 37.5% were positive for Cryptosporidium coproantigens. The highest rate of infection (78.1%) was observed in calves up to 3 ...
Background: AIDS and Protein energy malnutrition (PEM) severely impair the immune system Cryptosporidium has over the last two decades emerged as a life threatening disease. The study attempts to determine the prevalence of Cryptosporidium infection in malnourished children with HIV/AIDS. Method: Blood and stool ...
Holubová, Nikola; Sak, Bohumil; Horčičková, Michaela; Hlásková, Lenka; Květoňová, Dana; Menchaca, S.; McEvoy, J.; Kváč, Martin
Roč. 115, č. 6 (2016), s. 2243-2251 ISSN 0932-0113 R&D Projects: GA ČR GA15-01090S Institutional support: RVO:60077344 Keywords : Cryptosporidium avium * morphology * molecular analyses * transmission studies * Cryptosporidium avian genotype V Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 2.329, year: 2016
Enemark, Heidi L.; Ahrens, Peter; Bille-Hansen, Vivi
Genetic studies have demonstrated profound differences between the 'porcine' genotype of Cryptosporidium parvum, versus 'human' and 'bovine' genotypes. The study analysed infectivity and pathogenicity of the 'porcine' genotype (CPP-13 isolate) of C. parvum, and compared the results with published...... evidence for the existence of a new species of Cryptosporidium adapted to pigs....
The prevalence of the diarrhoea disease caused by the water-borne pathogens Cryptosporidium and Giardia in KwaZulu-Natal, was determined from pathology laboratory data. Cryptosporidium and Giardia were found to be endemic in KwaZulu-Natal with laboratory-confirmed incidences ranging from 2.9 to 3.7% and 2.9 to ...
Cryptosporidium species and cattle: Implication for public health and water - Short Communication. VA Pam, COE Onwuliri, DA Dakul, ICJ Omalu. Abstract. This paper presents a brief summary of the ecology of Cryptosporidium species in Calves and humans and the existing scientific evidence that addresses the claim that ...
Cryptosporidium and Cryptosporidiosis in Calves at Jos, Northern Nigeria. VA Pam, DA Dakul, COE Onwuliri. Abstract. This study investigated the occurrence of cryptosporidium and cryptosporidiosis in calves from Jos, Northern Nigeria. Two hundred fecal samples were collected from the calves, recruited for an all year ...
Jenkins, Mark C; Parker, Carolyn; Ritter, Donald
The purpose of this study was to determine if Eimeria oocyst concentrations and species composition in commercial broiler house litter changed during different cycles of anticoccidial drug (ACD) or live Eimeria oocyst vaccine (VAC) control programs and if there was a correlation between Eimeria oocyst levels and broiler performance. Litter samples were collected from a total of 15 different broiler farms encompassing a total of 45 individual houses during at least one complete grow-out cycle over a 21-mo period. Of these 15 broiler farms, three were followed for the entire 21-mo period spanning three ACD and four VAC cycles. Samples were collected at 2, 4, and 7-8 wk of grow-out corresponding to starter, grower, and withdraw periods of the ACD cycle. On a number of occasions, litter samples were obtained just prior to chick placement. Eimeria oocysts were isolated from all samples, counted by microscopy, and extracted for DNA to identify Eimeria species by ITS1 PCR. In general, Eimeria oocyst concentration in litter reached peak levels at 2-4 wk of grow-out regardless of coccidiosis control measure being used. However, peak oocyst numbers were sometimes delayed until 7-8 wk, indicating some level of Eimeria spp. drug resistance or incomplete vaccine coverage. Eimeria maxima , Eimeria acervulina , Eimeria praecox, and Eimeria tenella were generally present in all samples, and no difference in the species composition was noted between houses on a particular farm. While Eimeria species composition was similar among houses, Eimeria spp. oocyst levels exhibited sporadic peaks in one house of a given location's houses. Of particular interest was the observed correlation between E. maxima oocyst abundance and chick mortality. However, no correlation was observed in E. maxima oocyst levels, and the performance parameters adjusted feed conversion ratio and average daily weight gain. This study showed that understanding the dynamics of Eimeria spp. oocyst levels and species
Barta, John R; Thompson, R C Andrew
In raising the question "What is Cryptosporidium?", we aim to emphasize a growing need to re-evaluate the affinities of Cryptosporidium species within the phylum Apicomplexa so as to better understand the biology and ecology of these parasites. Here, we have compiled evidence from a variety of molecular and biological studies to build a convincing case for distancing Cryptosporidium species from the coccidia conceptually, biologically and taxonomically. We suggest that Cryptosporidium species must no longer be considered unusual or unique coccidia but rather seen for what they are--a distantly related lineage of apicomplexan parasites that are not in fact coccidia but that do occupy many of the same ecological niches. Looking at Cryptosporidium species without traditional coccidian blinders is likely to reveal new avenues of investigation into pathogenesis, epidemiology, treatment and control of these ubiquitous pathogens.
Xiao, Lihua; Escalante, Lillian; Yang, Chunfu; Sulaiman, Irshad; Escalante, Anannias A.; Montali, Richard J.; Fayer, Ronald; Lal, Altaf A.
Biological data support the hypothesis that there are multiple species in the genus Cryptosporidium, but a recent analysis of the available genetic data suggested that there is insufficient evidence for species differentiation. In order to resolve the controversy in the taxonomy of this parasite genus, we characterized the small-subunit rRNA genes of Cryptosporidium parvum, Cryptosporidium baileyi, Cryptosporidium muris, and Cryptosporidium serpentis and performed a phylogenetic analysis of t...
Chelladurai, J.J.; Clark, M.E.; Kváč, Martin; Holubová, Nikola; Khan, E.; Stenger, B.L.S.; Giddings, C.W.; McEvoy, J.
Roč. 115, č. 5 (2016), s. 1901-1906 ISSN 0932-0113 Institutional support: RVO:60077344 Keywords : Cryptosporidium * Red-winged blackbird * Passerines * Cryptosporidium galli * Avian genotypeVI * Proventriculus * Intestine Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 2.329, year: 2016
Němejc, K.; Sak, Bohumil; Květoňová, Dana; Hanzal, V.; Jeníková, Martina; Kváč, Martin
Roč. 184, 2/4 (2012), 122-125 ISSN 0304-4017 Grant - others:Mšk(CZ) 6007665806 Institutional research plan: CEZ:AV0Z60220518; CEZ:AV0Z50450515 Keywords : Cryptosporidium suis * Cryptosporidium pig genotype II * Eurasian wild boar * SSU * PCR Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 2.381, year: 2012
Jian, Fuchun; Liu, Aiqin; Wang, Rongjun; Zhang, Sumei; Qi, Meng; Zhao, Wei; Shi, Yadong; Wang, Jianling; Wei, Jiujian; Zhang, Longxian; Xiao, Lihua
Extensive genetic variation is observed within the genus Cryptosporidium and the distribution of Cryptosporidium species/genotypes in humans and animals appears to vary by geography and host species. To better understand the genetic diversity of Cryptosporidium spp. in horses and donkeys, we characterized five horse-derived and 82 donkey-derived Cryptosporidium isolates from five provinces or autonomous regions (Sichuan, Gansu, Henan, Inner Mongolia and Shandong) in China at the species/genotype and subtype levels. Three Cryptosporidium species/genotypes were identified based on the analysis of the SSU rRNA gene, including Cryptosporidium parvum (n=22), the Cryptosporidium horse genotype (n=4), and Cryptosporidium hominis (n=61). The identification of C. hominis was confirmed by sequence analysis of the HSP70 and actin genes. Subtyping using sequence analysis of the 60kDa glycoprotein gene identified 21 C. parvum isolates as subtype IIdA19G1, the four horse genotype isolates as subtypes VIaA15G4 (n=2) and VIaA11G3 (n=2), and the 61 C. hominis isolates as IkA16G1 (n=59) and IkA16 (n=2). The common finding of C. hominis reaffirms the heterogeneity of Cryptosporidium spp. in horses and donkeys and is possibly a reflection of endemic transmission of C. hominis in these animals. Data of the study suggest that horses and donkeys as companion animals may potentially transmit Cryptosporidium infections to humans. Copyright © 2016 Elsevier B.V. All rights reserved.
Chalmers, Rachel M; Elwin, Kristin; Cheesbrough, John; Hadfield, Stephen J; Beeching, Nicholas J
Detection of anti-Cryptosporidium immunoglobulin G (IgG) antibodies in human sera has been used to demonstrate population exposure to this gastro-intestinal protozoan parasite. We characterised the dynamics of IgG antibody responses to two Cryptosporidium parvum (IOWA isolate) sporozoite antigens (15/17 kDa and 27 kDa) using longitudinal sera taken from laboratory-confirmed cryptosporidiosis cases in England and Wales. The effect of the infecting Cryptosporidium species was also investigated. A mini-gel Western blot was used to test sera from ten Cryptosporidium stool-positive diarrhoea patients, taken soon after diagnosis and at 3 month intervals. Overall responses to the 15/17 kDa antigen complex were stronger and over a greater range than those to the 27 kDa antigen, but declined between 181 and 240 days and were barely detectable thereafter. Responses to the 27 kDa antigen were much weaker but remained detectable for a greater length of time. No differences were detected in either antibody response to infection with C. hominis or C. parvum. The assay appears to be applicable for the study of recent exposure to C. parvum or C. hominis in the United Kingdom population, with strong responses to the 15/17 kDa antigen occurring within 6 months of infection. Copyright © 2013 The British Infection Association. Published by Elsevier Ltd. All rights reserved.
Subtype Analysis of Cryptosporidium Specimens from Sporadic Cases in Colorado, Idaho, New Mexico, and Iowa in 2007: Widespread Occurrence of One Cryptosporidium hominis Subtype and Case History of an Infection with the Cryptosporidium Horse Genotype▿
Xiao, Lihua; Hlavsa, Michele C.; Yoder, Jonathan; Ewers, Christina; Dearen, Theresa; Yang, Wenli; Nett, Randall; Harris, Stephanie; Brend, Sarah M.; Harris, Meghan; Onischuk, Lisa; Valderrama, Amy L.; Cosgrove, Shaun; Xavier, Karen; Hall, Nancy
Subtyping was conducted in late 2007 on 57 Cryptosporidium specimens from sporadic cases in Colorado, Idaho, New Mexico, and Iowa. One previously rare Cryptosporidium hominis subtype was indentified in 40 cases (70%) from all four states, and the Cryptosporidium horse genotype was identified in a pet shop employee with severe clinical symptoms.
Subtype analysis of Cryptosporidium specimens from sporadic cases in Colorado, Idaho, New Mexico, and Iowa in 2007: widespread occurrence of one Cryptosporidium hominis subtype and case history of an infection with the Cryptosporidium horse genotype.
Xiao, Lihua; Hlavsa, Michele C; Yoder, Jonathan; Ewers, Christina; Dearen, Theresa; Yang, Wenli; Nett, Randall; Harris, Stephanie; Brend, Sarah M; Harris, Meghan; Onischuk, Lisa; Valderrama, Amy L; Cosgrove, Shaun; Xavier, Karen; Hall, Nancy; Romero, Sylvia; Young, Stephen; Johnston, Stephanie P; Arrowood, Michael; Roy, Sharon; Beach, Michael J
Subtyping was conducted in late 2007 on 57 Cryptosporidium specimens from sporadic cases in Colorado, Idaho, New Mexico, and Iowa. One previously rare Cryptosporidium hominis subtype was identified in 40 cases (70%) from all four states, and the Cryptosporidium horse genotype was identified in a pet shop employee with severe clinical symptoms.
Jeníková, M.; Němejc, K.; Sak, Bohumil; Květoňová, Dana; Kváč, Martin
Roč. 176, 2/3 (2011), 120-125 ISSN 0304-4017 R&D Projects: GA ČR GP523/07/P117 Institutional research plan: CEZ:AV0Z60220518 Keywords : Cryptosporidium suis * Cryptosporidium pig genotype II * Mixed infection * Age-specificity * Species-specific primers Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 2.579, year: 2011
Uzeda, Rosangela Soares; Costa, Kattyanne De Souza; Santos, Sara Lima; Pinheiro, Alexandre Moraes; De Almeida, Maria Angela Ornelas; McAllister, Milton M.
The purpose of this study was to investigate whether sporulated Neospora caninum oocysts, which had been stored for 46 mo in a 2% sulfuric acid solution at 4℃, remain morphologically viable and infective to gerbils (Meriones unguiculatus). Six gerbils were orally inoculated with doses of 400 or 1,200 oocysts. Two mo after inoculation, the animals did not show any clinical signs, had no histological lesions, and were seronegative for N. caninum at 1: 50 in an immunofluorescent antibody test. PCR using the brain from each gerbil did not reveal N. caninum specific DNA. We conclude that oocysts preserved for 46 mo are not infective, despite being morphologically intact. PMID:18165712
Yan, Wenchao; Alderisio, Kerri; Roellig, Dawn M; Elwin, Kristin; Chalmers, Rachel M; Yang, Fengkun; Wang, Yuanfei; Feng, Yaoyu; Xiao, Lihua
Cryptosporidium skunk genotype is a zoonotic pathogen commonly identified in surface water. Thus far, no subtyping tool exists for characterizing its transmission in humans and animals and transport in environment. In this study, a subtyping tool based on the 60kDa glycoprotein (gp60) gene previously developed for Cryptosporidium chipmunk genotype I was used in the characterization of Cryptosporidium skunk genotype in animal and storm runoff samples from a watershed in New York. Altogether, 17 positive samples from this watershed and 5 human and animal specimens from other areas were analyzed. We identified 14 subtypes of Cryptosporidium skunk genotype, 11 of which were seen in the watershed. In phylogenetic analysis, these subtypes belonged to 4 subtype families (XVIa, XVIb, XVIc, and XVId). No host-adapted subtypes were identified and the two subtypes in humans were genetically similar to some in raccoons, otters, and storm runoff samples from the watershed. The characteristics of gp60 protein sequences of the Cryptosporidium skunk genotype are similar to those of other Cryptosporidium species, but only its XVIb subtype family has a putative furin cleavage site. This subtyping tool might be useful in characterizing Cryptosporidium skunk genotype in clinical and environmental samples. Copyright © 2017 Elsevier B.V. All rights reserved.
Blanco, María Alejandra; Iborra, Asunción; Vargas, Antonio; Nsie, Eugenia; Mbá, Luciano; Fuentes, Isabel
The aim of the study was to perform a molecular characterization of clinical isolates of Cryptosporidium species from Equatorial Guinea. Standard laboratory methods were used to identify 35 cryptosporidiosis cases among 185 patients. PCR-RFLP successfully identified 34 Cryptosporidium species from these 35 cases, comprising C. parvum (52.9%), C. hominis (44.1%) and C. meleagridis (2.9%); over 90% of the species were isolated from HIV-positive patients. This is the first report of the molecular characterization of Cryptosporidium species isolated from humans in Equatorial Guinea and shows that zoonotic and anthroponotic transmission is present in this country.
Prevalência de Cryptosporidium spp e Giardia sp em eqüinos estabulados no Jockey Club de Santa Maria - RS, Brasil Prevalence of Cryptosporidium spp. and Giardia sp. infection in horses stabled in the Jockey Club of Santa Maria - RS, Brazil
Aline Diefenbach Gomes
Full Text Available O Cryptosporidium spp. e a Giardia sp. são atualmente reconhecidos como os principais patógenos entéricos com potencial zoonótico. O presente estudo visou estabelecer a prevalência desses protozoários em eqüinos hospedados no Jockey Club de Santa Maria, RS, Brasil, no período de 19 de maio a 30 de junho de 2007. Foram coletadas amostras de fezes, diretamente da ampola retal, de 64 animais. As amostras de fezes foram processadas por meio do método de centrifugação-flutuação de Faust modificado. Posteriormente essas amostras foram visualizadas ao microscópio óptico para a pesquisa de cistos e oocistos. Os resultados encontrados revelaram a presença de Cryptosporidium spp. em 75% (48/64 das amostras. Cistos de Giardia sp. não foram encontrados nas amostras de fezes analisadas. A freqüência de Cryptosporidium spp. nas diferentes faixas etárias foi de 83,3% (15/18 nos potros até dois anos de idade, 71% (22/31 nos jovens entre dois e cinco anos e 80% (12/15 nos adultos. Os resultados demonstram que o Cryptosporidium spp. está amplamente disseminado na população de eqüinos do Jockey Club de Santa Maria e pode representar uma fonte de infecção significativa para a população da região.Cryptosporidium spp. and Giardia sp. are currently recognised as the main enteric pathogens with potential zoonotic transmission risk. The present study aimed to investigate the prevalence of these parasites in horses stabled in the Santa Maria Jockey Club between May 19 and June 30, 2007. Fecal samples from 64 horses were collected directly from the animals’ rectal ampoule. The 64 fecal samples were processed using modified Faust’s method through the centrifugation-floatation technique, and were then later visualized under optical microscope for detection of Cryptosporidium spp. oocysts and Giardia sp. cysts. The results showed the occurrence of Cryptosporidium spp. in 75% (48/64 of the samples. Giardia sp. cysts were not found in the
Al Mawly, Julanda; Grinberg, Alex; Velathanthiri, Niluka; French, Nigel
The estimation of the prevalence and zoonotic potential of Cryptosporidium parvum cycling in bovine populations requires the use of genotyping, as several morphologically similar non-parvum genetic variants of unproven clinical and public health impact are found in cattle. However, robust C. parvum prevalence estimates in cattle are lacking and comparative data of bovine and human isolates collected from the same regions are scarce. Thus, the relative contribution of the C. parvum oocysts released by farmed animals to animal and human cryptosporidiosis burden is, in general, poorly understood. The New Zealand farm-level C. parvum prevalence was estimated using a cross-sectional sample of 1283 faecal specimens collected from newborn calves on 97 dairy farms. Faeces were analysed by immunofluorescence and the Cryptosporidium parasites were genetically identified. Finally, bovine C. parvum were genetically compared with historical human clinical isolates using a bilocus subtyping scheme. Immunofluoresence-positive faeces were found in 63/97 (65%) farms. C. parvum was identified in 49 (50.5%) farms, C. bovis in 6 (6.1%) farms, and on 8 (8.2%) farms the species could not be identified. The dominant C. parvum genetic variants were geographically widespread and found in both host populations, but several variants were found in humans only. Phenotypic tests offered by New Zealand veterinary diagnostic laboratories for the diagnosis of C. parvum may have moderate to high positive predictive values for this species. The genetic similarities observed between the human and bovine parasites support a model considering calves as significant amplifiers of zoonotic C. parvum in New Zealand. However, data suggest that transmission routes not associated with dairy cattle should also be taken into account in future source-attribution studies of human cryptosporidiosis.
Zhao, Wei; Wang, Rongjun; Zhang, Weizhe; Liu, Aiqin; Cao, Jianping; Shen, Yujuan; Yang, Fengkun; Zhang, Longxian
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. PMID:24999982
John J. Debenham
Full Text Available Giardia duodenalis, Cryptosporidium spp., and Entamoeba spp. are intestinal protozoa capable of infecting a range of host species, and are important causes of human morbidity and mortality. Understanding their epidemiology is important, both for public health and for the health of the animals they infect. This study investigated the occurrence of these protozoans in rhesus macaques (Macaca mulatta in India, with the aim of providing preliminary information on the potential for transmission of these pathogens between macaques and humans. Faecal samples (n = 170 were collected from rhesus macaques from four districts of North-West India. Samples were analysed for Giardia/Cryptosporidium using a commercially available direct immunofluorescent antibody test after purification via immunomagnetic separation. Positive samples were characterised by sequencing of PCR products. Occurrence of Entamoeba was investigated first by using a genus-specific PCR, and positive samples further investigated via species-specific PCRs for Entamoeba coli, Entamoeba histolytica, Entamoeba dispar and Entamoeba moshkovskii. Giardia cysts were found in 31% of macaque samples, with all isolates belonging to Assemblage B. Cryptosporidium oocysts were found in 1 sample, however this sample did not result in amplification by PCR. Entamoeba spp. were found in 79% of samples, 49% of which were positive for E. coli. Multiplex PCR for E. histolytica, E. dispar and E. moshkovskii, did not result in amplification in any of the samples. Thus in 51% of the samples positive at the genus specific PCR, the Entamoeba species was not identified. This study provides baseline information on the potential for transmission of these zoonotic parasites at the wildlife-human interface.
, 2006, South Africa ... applied in the United Kingdom and New. Zealand (DWI, 2008; NZ Ministry of Health, 2008). In South. Africa, Cryptosporidium and Giardia monitoring in drink- ...... Improvement plans implemented.
Hong, Semie; Kim, Kyungjin; Yoon, Sejoung; Park, Woo-Yoon; Sim, Seobo; Yu, Jae-Ran
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.
Yang, Wenli; Chen, Plato; Villegas, Eric N.; Landy, Ronald B.; Kanetsky, Charles; Cama, Vitaliano; Dearen, Theresa; Schultz, Cherie L.; Orndorff, Kenneth G.; Prelewicz, Gregory J.; Brown, Miranda H.; Young, Kim Roy; Xiao, Lihua
To better characterize Cryptosporidium in the Potomac River watershed, a PCR-based genotyping tool was used to analyze 64 base flow and 28 storm flow samples from five sites in the watershed. These sites included two water treatment plant intakes, as well as three upstream sites, each associated with a different type of land use. The uses, including urban wastewater, agricultural (cattle) wastewater, and wildlife, posed different risks in terms of the potential contribution of Cryptosporidium...
Takumi, Katsuhisa; Cacci?, Simone M.; van der Giessen, Joke; Xiao, Lihua; Sprong, Hein
Background Cryptosporidiosis is a gastrointestinal disease affecting many people worldwide. Disease incidence is often unknown and surveillance of human cryptosporidiosis is installed in only a handful of developed countries. A genetic marker that mirrors disease incidence is potentially a powerful tool for monitoring the two primary human infected species of Cryptosporidium. Methods We used the molecular epidemiological database with Cryptosporidium isolates from ZoopNet, which currently con...
Besasso, H R; Macías, J; Trombetta, L A; di Risio, C A; Argento, R; Besuschio, S C
Occurrence of vomiting and diarrhea in patients with acquired immunodeficiency syndrome was most frequently attributed to enteropathogens organism that invade the gastrointestinal tract because of the immunologic unbalance of the host... Among several causes the cryptosporidium has been detected quite often and its predominant localization was the gastrointestinal tract, although other extraintestinal sites has also been reported. In both cases, erosive congestive gastritis was found, while histological examination showed cryptosporidium in gastric biopsy specimens.
Schiller, Sabine Eva; Webster, Koa Narelle; Power, Michelle
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
Qi, Meng; Wang, Rongjun; Jing, Bo; Jian, Fuchun; Ning, Changshen; Zhang, Longxian
Cryptosporidium andersoni is the predominant species in post-weaned and adult cattle in China. The aim of this study was to determine the prevalence and understand the transmission of cattle cryptosporidiosis in the Xinjiang Uyghur Autonomous Region, China, a total of 1827 fecal samples (436 from He cattle and 1391 from dairy cattle) were examined for the presence of C. andersoni-like oocysts by microscopy after Sheather's sugar flotation technique. The overall prevalence of C. andersoni-like was 3.8% (70/1827) and all the C. andersoni-like isolates were identified as C. andersoni at the SSU rRNA locus. Among the C. andersoni isolates, a total of 60 isolates were successfully characterized into eight multilocus sequence typing (MLST) subtypes using MLST analysis at the four microsatellite/minisatellite loci (MS1, MS2, MS3 and MS16), and three new subtypes were identified. The MLST subtype A4,A4,A4,A1 showed a predominance and a wide distribution among the eight MLST subtypes obtained in the investigated areas. The MLST subtypes A2,A4,A2,A1 and A4,A5,A2,A1 showed a unique distribution in the investigated areas. A linkage disequilibrium analysis showed the presence of an epidemic population genetic structure of C. andersoni isolated from dairy and He cattle in Xinjiang. These findings provide new insights into the genetic structure of C. andersoni isolates and are also helpful to explore the infection source of C. andersoni in cattle in Xinjiang, China. Copyright © 2016 Elsevier B.V. All rights reserved.
Codices, Vera; Martins, Catarina; Novo, Carlos; Pinho, Mário; de Sousa, Bruno; Lopes, Angela; Borrego, Miguel; Matos, Olga
Cryptosporidium parvum is an intracellular parasite causing enteritis which can become life-threatening in immunocompromised host. Immunoregulatory T cells play a central role in the regulatory network of the host. Here, we proposed to characterize the populations of immune cells during infection and reinfection with C. parvum. Four-week-old BALB/C mice were inoculated with oocysts of C. parvum at days 0 and 22. Fecal and blood samples, spleens, and small intestines were collected for analysis. Peripheral blood and spleen cell populations were characterized by flow cytometry. After infection (days 0 to 21), mice presented higher values of neutrophils, eosinophils, NK cells and CD4(+)CD25(high) T cells in peripheral blood. After reinfection, this upward trend continued in the following days for all four populations in infected mice. At day 35, infected mice presented similar values to the control group, except for CD4(+)CD25(high) T cells, which remained higher in infected mice. A possible correlation between alterations in blood and spleen cell populations was also studied, but no consistent association could be established. Small intestine sections were screened for intracellular stages of the parasite but no evidence of pathology was observed. Here, we report information which may be important for the understanding of the specific cell-mediated response in immunocompetent mice to C. parvum infection. Although some questions remain unanswered and complementary studies are needed, our results are expected to contribute to a better understanding of innate and Treg cells role in the clearance process of this parasite.
Djokic, Vitomir; Blaga, Radu; Rinaldi, Laura; Le Roux, Delphine; Ducry, Tamara; Maurelli, Maria Paola; Perret, Catherine; Djurkovic Djakovic, Olgica; Cringoli, Giuseppe; Boireau, Pascal
Oocysts of Toxoplasma gondii represent one of the most common environmental contaminants causing the zoonotic infection toxoplasmosis. The aim of the present study was to compare the Mini-FLOTAC device with traditional cell counting plates (Kova Slide) for the detection of T. gondii oocysts from feline feces. Two types of experiments were performed: (i) purified oocysts were counted in different dilutions and (ii) specific pathogen free T. gondii-negative cat feces was inoculated with numbers of purified oocysts and counting was performed directly from feces. Our analysis showed a thousand times higher sensitivity of Mini-FLOTAC (5 × 10(2) oocysts) compared to Kova Slide (5 × 10(5) oocysts). Also, when compared by McNemar's test, counting of the purified oocysts showed a higher sensitivity of Mini-FLOTAC compared to Kova Slide, for a dilution of 10(3) oocysts/ml (chi(2) = 6.1; P microscopes in any laboratory or field conditions. We therefore recommend its use for regular screening. Further studies are needed to validate Mini-FLOTAC for the detection of oocysts in soil and water samples in field conditions. Copyright © 2014 Elsevier Inc. All rights reserved.
Liu, Aiqin; Zhang, Jia; Zhao, Jingmin; Zhao, Wei; Wang, Rongjun; Zhang, Longxian
Background 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...
Wang, Tao; Chen, Zuqin; Xie, Yue; Hou, Rong; Wu, Qidun; Gu, Xiaobing; Lai, Weiming; Peng, Xuerong; Yang, Guangyou
Background 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. Findin...
Youn, Sojin; Kabir, Mamun; Haque, Rashidul; Petri, William A.
The Giardia/Cryptosporidium Chek test (TechLab, Inc.), a screening test for Giardia and Cryptosporidium, was evaluated with 136 fecal samples. Using the results of the Giardia II test and Cryptosporidium II test as gold standards, it was 98.4% sensitive and 100% specific and had positive and negative predictive values of 98.7% and 99.3%.
de Lucio, Aida; Bailo, Begoña; Aguilera, María; Cardona, Guillermo A; Fernández-Crespo, Juan C; Carmena, David
The role of pet dogs and cats as suitable source of human infections by the diarrheagenic protozoan parasites Giardia duodenalis and Cryptosporidium spp. has been a topic of intense debate for long time and still remains a largely unsolved problem. In this cross-sectional molecular epidemiological survey we attempted to investigate whether zoonotic (or zooanthroponotic) disease transmission was occurring among humans and domestic dogs and cats sharing the same spatial and temporal setting in both rural and urban areas of the province of Álava, Northern Spain. A total of 268 (including 179 human, 55 canine, and 34 feline) individual faecal specimens were obtained from 63 family households during February-March and November-December 2014. Detection of G. duodenalis cysts and Cryptosporidium spp. oocysts was achieved by direct fluorescence microscopy (DFAT) and PCR-based methods targeting the small subunit (SSU) ribosomal RNA gene of the parasites. Giardia-positive isolates were subsequently sub-genotyped at the glutamate dehydrogenase (GDH) and β-giardin (BG) genes. Overall, G. duodenalis infections were identified in 3.4% (6/179) of humans, 29% (16/55) of dogs, and 5.9% (2/34) of cats, respectively. Cryptosporidium spp. infections were detected in 1.1% (2/179) of humans, 5.5% (3/55) of dogs, and 8.8% (3/34) of cats, respectively. Simultaneous infections in human and canine/feline hosts by G. duodenalis or Cryptosporidium spp. were only demonstrated in a single household in which a cat and its owner tested positive for Cryptosporidium by DFAT, but this result could not be confirmed by SSU-PCR. Infections were homogeneously distributed among the studied human or animal populations irrespectively of their sex, age group, or geographical region of origin. Inadequate washing of raw vegetables and fruits was the only risk factor significantly associated to a higher likelihood of having human giardiosis/cryptosporidiosis. Molecular characterization of G. duodenalis
Full Text Available The in vitro production of gametocytes and oocysts of the apicomplexan parasite genus Eimeria is still a challenge in coccidiosis research. Until today, an in vitro development of gametocytes or oocysts had only been shown in some Eimeria species. For several mammalian Eimeria species, partial developments could be achieved in different cell types, but a development up to gametocytes or oocysts is still lacking. This study compares several permanent cell lines with primary fetal cells of the black rat (Rattus norvegicus concerning the qualitative in vitro development of the rat parasite Eimeria nieschulzi. With the help of transgenic parasites, the developmental progress was documented. The selected Eimeria nieschulzi strain constitutively expresses the yellow fluorescent protein and a macrogamont specific upregulated red tandem dimer tomato. In the majority of all investigated host cells the development stopped at the second merozoite stage. In a mixed culture of cells derived from inner fetal organs the development of schizont generations I-IV, macrogamonts, and oocysts were observed in crypt-like organoid structures. Microgamonts and microgametes could not be observed and oocysts did not sporulate under air supply. By immunohistology, we could confirm that wild-type E. nieschulzi stages can be found in the crypts of the small intestine. The results of this study may be helpful for characterization of native host cells and for development of an in vitro cultivation system for Eimeria species.
Helmy, Yosra A; Krücken, Jürgen; Nöckler, Karsten; von Samson-Himmelstjerna, Georg; Zessin, Karl-H
For the detection of Cryptosporidium species in 804 animals and 165 diarrhoeic children (tests, the RIDASCREEN® Cryptosporidium test [enzyme immunoassay (EIA)] and the RIDA®QUICK Cryptosporidium/Giardia Combi [immuno-chromatographic test (ICT)] as well as polymerase chain reaction (PCR) were used. Prevalence of Cryptosporidium was 15.0, 19.5 and 32.3% in animals and 2.4, 6.7 and 49.1% in children using EIA, ICT and PCR, respectively.Using PCR as reference method, animal samples sensitivity (Se) of the EIA was 46.5% when questionable samples were considered positive, whereas specificity (Sp) was 100%. Se of the ICT was 60.4% while Sp was 100%. Positive predictive values (PPVs) for both EIA and ICT test were 100%, and negative predictive values (NPVs) for EIA were 79.7 and 84.1% for ICT. For the children samples, the Se of EIA was 5%, Sp was 100%, PPV was 100% and NPV was 52.2%, while the Se of ICT was 13.6%, Sp was 100%, PPV was 100% and NPV was 54.6%.The Kappa score of agreement between PCR and ICT was 67.4%, 54.1% between PCR and EIA and 84.4% between ICT and EIA. Until the second serial dilution of the EIA and ICT test, 9 × 10(3) oocysts/μl of Cryptosporidia was detected, whereas in PCR, they were detected until the sixth serial dilution. Copro-antigen tests were easy to perform and less time-consuming but less sensitive compared to PCR. They obviously are best applicable for screening and epidemiological studies of large numbers of subjects, for batch specimen processing and in isolated or rural areas where reliable tests like PCR are unfeasible. When in children, a single stool sample is used for the diagnosis of clinical cases; better results can be obtained when non-standardized PCR due low specificity is coupled with copro-antigen tests.
Ifeonu, Olukemi O; Simon, Raphael; Tennant, Sharon M; Sheoran, Abhineet S; Daly, Maria C; Felix, Victor; Kissinger, Jessica C; Widmer, Giovanni; Levine, Myron M; Tzipori, Saul; Silva, Joana C
Human cryptosporidiosis, caused primarily by Cryptosporidium hominis and a subset of Cryptosporidium parvum, is a major cause of moderate-to-severe diarrhea in children under 5 years of age in developing countries and can lead to nutritional stunting and death. Cryptosporidiosis is particularly severe and potentially lethal in immunocompromised hosts. Biological and technical challenges have impeded traditional vaccinology approaches to identify novel targets for the development of vaccines against C. hominis, the predominant species associated with human disease. We deemed that the existence of genomic resources for multiple species in the genus, including a much-improved genome assembly and annotation for C. hominis, makes a reverse vaccinology approach feasible. To this end, we sought to generate a searchable online resource, termed C. hominis gene catalog, which registers all C. hominis genes and their properties relevant for the identification and prioritization of candidate vaccine antigens, including physical attributes, properties related to antigenic potential and expression data. Using bioinformatic approaches, we identified ∼400 C. hominis genes containing properties typical of surface-exposed antigens, such as predicted glycosylphosphatidylinositol (GPI)-anchor motifs, multiple transmembrane motifs and/or signal peptides targeting the encoded protein to the secretory pathway. This set can be narrowed further, e.g. by focusing on potential GPI-anchored proteins lacking homologs in the human genome, but with homologs in the other Cryptosporidium species for which genomic data are available, and with low amino acid polymorphism. Additional selection criteria related to recombinant expression and purification include minimizing predicted post-translation modifications and potential disulfide bonds. Forty proteins satisfying these criteria were selected from 3745 proteins in the updated C. hominis annotation. The immunogenic potential of a few of these is
Yang, Rongchang; Fenwick, Stanley; Potter, Abbey; Ng, Josephine; Ryan, Una
A total of 763 faecal samples were collected from western grey kangaroos (Macropus fuliginosus) in Western Australia and screened for the presence of Cryptosporidium by PCR at the 18S ribosomal RNA (rRNA) locus. Samples that were positive at the 18S locus were also amplified at the actin locus. The overall prevalence was 9.3% (71/763). At the 18S rRNA locus, sequences were obtained for 28 of the 71 positives. Sequence analysis identified four species; Cryptosporidium fayeri in seven isolates, Cryptosporidium marcopodum in four isolates, Cryptosporidium xiaoi in six isolates and a novel genotype (kangaroo genotype I) in eleven isolates. Analysis at the actin locus confirmed the genetic distinctness of the novel genotype. The results of the present study indicate that in addition to C. fayeri and C. marcopodum, kangaroos may be capable of being infected with a wider range of Cryptosporidium species and genotypes including livestock species such as C. xiaoi. The novel genotype identified in the kangaroos most likely represents a cryptic species that requires further analyses to confirm its species status. Copyright © 2011 Elsevier B.V. All rights reserved.
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
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. © C. Gong et al., published by EDP Sciences, 2017.
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.
Al-Mekhlafi, H M; Mahdy, M A K; ’Azlin, M Y; Fatmah, M S; Norhayati, M
Cryptosporidium is a coccidian parasite that is prevalent worldwide, some species of which cause morbidity in both immunocompromised and immunocompetent individuals. The prevalence and predictors of Cryptosporidium infection, and its effect on nutritional status, have recently been explored among 276 children (141 boys and 135 girls, aged 2–15 years) in aboriginal (Orang Asli) villages in the Malaysian state of Selangor. Faecal smears were examined by the modified Ziehl–Neelsen staining technique while socio–economic data were collected using a standardized questionnaire. Nutritional status was assessed by anthropometric measurements. Cryptosporidium infection, which was detected in 7·2% of the aboriginal children, was found to be significantly associated with low birthweight (⩽2·5 kg), being part of a large household (with more than seven members) and prolonged breast feeding (>2 years). The output of a binary logistic regression confirmed that large household size was a significant predictor of Cryptosporidium infection (giving an odds ratio of 2·15, with a 95% confidence interval of 1·25–5·02). Cryptosporidium infection is clearly a public-health problem among the aboriginal children of Selangor, with person-to-person the most likely mode of transmission. PMID:21396250
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.
Wagnerová, Pavla; Sak, Bohumil; McEvoy, J.; Rost, M.; Perec Matysiak, A.; Ježková, J.; Kváč, Martin
Roč. 114, č. 4 (2015), s. 1619-1624 ISSN 0932-0113 R&D Projects: GA ČR GA15-01090S Institutional support: RVO:60077344 Keywords : horse * Cryptosporidium * SSU * gp60 * MLST Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 2.027, year: 2015
Dubey, J. P.; Andrews, C.D.; Lind, Peter
Objective-To follow antibody responses measured by various serologic tests in pigs orally inoculated with low (less than or equal to 10 oocysts) numbers of Toxoplasma gondii oocysts. Animals-24, 2- to 3-month-old pigs. Procedure-Pigs (n = 42) were inoculated orally with 10 (14 pigs) or 1 (28 pigs...
by multivariate statistical techniques. The morphology of 810 Eimeria specimens was defined in binary (b/w) digital images by pixels of their oocyst outline. A Fourier transform of pixel positions yielded size and shape features. To classify coccidia, the quantitative data were employed in an agglomerative...... clustering by average linkage algorithm with equal weight assigned to size and shape. An inverse Fourier transform served to reconstruct oocyst outlines, i.e. outlines of average shape and size, from mean values of features in resulting clusters. Clusters were subsequently identified based on their average...... after reconstruction by inverse Fourier transform and comparison with earlier studies....
Presencia de Giardia lamblia y Cryptosporidium spp. en aguas residuales depuradas reutilizadas para riego agrícola en la isla de Tenerife, España. Efectos del transporte a larga distancia sobre la calidad del agua reutilizada Giardia lamblia and Cryptosporidium spp. Presence in treated wastewater reutilised for irrigation in Tenerife island, Spain. Long-distance transport effects in the reutilised water quality
Escolástico Aguiar González
habitats. These protozoan investigation and detection have acquired importance in the last years due to their dispersion forms, which show resistance to the habitual treatments of potabilization and purification, and their classification as emerging pathogens which are the causative agents of important hydrical transmission outbreaks.In our work, Giardia lamblia cyst and Cryptosporidium spp. oocyst presence in Santa Cruz de Tenerife treated wastewater which is transported to the South of the island to be reutilised in agriculture. Furthermore, transport, storage and advanced treatment effects against cyst and oocyst concentration throughout the system and relation existence with other bacteriological and physical-chemical parameters, are also investigated.The obtained results demonstrate variable behaviours in cyst and oocyst concentration against applied treatments, and the depurative effect of the long-distance transport which seems to have in treated wastewater.There was not found any relation between cyst and oocyst concentration in treated wastewater and the traditional indicators of faecal contamination.
production. Proc. Soc. Exptl. Biol. Med. 116:174-177. Mayer, V. 1965. Study of the virulence of tick-borne encephalitis virus. IV. Thermosensitivity...inactivation of rabies and other rhabrtoviruses: stabilization of the chelating agent Ethylenediaminetetraacetic acid at physiological temperatures. Infec
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.
Tanrıverdi, Sultan; Grinberg, Alex; Chalmers, Rachel M.; Hunter, Paul R.; Petrovic, Zorana; Akiyoshi, Donna E.; London, Eric; Zhang, Linghui; Tzipori, Saul; Tumwine, James K.; Widmer, Giovanni
Cryptosporidium parvum and Cryptosporidium hominis are two related species of apicomplexan protozoa responsible for the majority of human cases of cryptosporidiosis. In spite of their considerable public health impact, little is known about the population structures of these species. In this study, a battery of C. parvum and C. hominis isolates from seven countries was genotyped using a nine-locus DNA subtyping scheme. To assess the existence of geographical partitions, the multilocus genotype data were mined using a cluster analysis based on the nearest-neighbor principle. Within each country, the population genetic structures were explored by combining diversity statistical tests, linkage disequilibrium, and eBURST analysis. For both parasite species, a quasi-complete phylogenetic segregation was observed among the countries. Cluster analysis accurately identified recently introduced isolates. Rather than conforming to a strict paradigm of either a clonal or a panmictic population structure, data are consistent with a flexible reproductive strategy characterized by the cooccurrence of both propagation patterns. The relative contribution of each pattern appears to vary between the regions, perhaps dependent on the prevailing ecological determinants of transmission. PMID:18836013
Due to its extensive polymorphism, a partial sequence of the Cryptosporidium surface glycoprotein gene gp60 has been frequently used as a genetic marker. I explored the global diversity of this protein, and compared its sequence diversity in Cryptosporidium parvum and Cryptosporidium hominis. In marked contrast to the geographical partition of C. parvum and C. hominis multi-locus genotypes, gp60 allelic groups showed no evidence of segregating in space, or of differing with respect to geographical diversity. Globally, genetic diversity of C. hominis gp60 exceeded that of C. parvum. Within C. parvum, gp60 alleles originating from human isolates were more diverse than those infecting ruminants. Phylogenetic analysis grouped gp60 sequences into a small number of relatively homogenous allelic groups, with only a small number of alleles having evolved independently. With the notable exception of a group of alleles restricted to humans, C. parvum alleles are found in ruminants and humans.
Dasgupta, T; Lee, E H
A patented gel delivery system being used to deliver coccidiosis vaccine to poultry hatchlings is assessed. For effective vaccination, the coccidial oocysts must be uniformly suspended before exposure to birds. The uniformity of distribution within the gel was evaluated by incorporating a culture of chicken gut flora into gel sausages, placing sections of the sausage on culture plates, determining the appearance and distribution of bacterial colonies on culture plates after incubation, and verifying by cell counts. The uniformity of distribution of similarly prepared coccidial oocysts was verified by infecting birds with 40,000 Eimeria tenella oocysts delivered via the gel. Gel-inoculated birds were compared with control birds inoculated PO with 40,000 oocysts suspended in water by using cecal lesion scores. Both the appearance and colony counts of chicken gut flora from the gel were uniform. The standard deviation in the lesion scores for the gel-inoculated group and the water-inoculated groups were 0.51 and 0.69, respectively. The results indicate that a gel delivery system can provide uniform distribution of live organisms and vaccine agents to birds. Images Figure 1a. Figure 1b. PMID:10945126
Enemark, Heidi L.; Dahl, Jan; Enemark, Jörg M Dehn
samples from dairy herds (n = 52) with > 50 cows and a history of diarrhoea in young stock. Individual faecal samples were collected 3–4 weeks following re-housing to common pens from calves (n = 453) aged 3 weeks to 6 months. Faecal consistency and total number of oocysts per gram of faeces (opg) were...
Detection of Cryptosporidium sp. in non diarrheal faeces from children, in a day care center in the city of São Paulo, Brazil Detecção de Cryptosporidium sp. em fezes não diarréicas de crianças, em uma escola de educação infantil de São Paulo, Brasil
Therezinha T. Carvalho-Almeida
Full Text Available The protozoan Cryptosporidium sp. has been frequently detected in faeces from children with persistent diarrhoea. This work achieved to investigate an outbreak of cryptosporidiosis, in a day care center, attending children of high socio-economic level, between 0 and six years old. The outbreak was detected through the network of public health, when stool samples, not diarrhoeic, were examined at the Parasitology Service of the Adolfo Lutz Institute. Among the 64 examined children, 13 (20.3% showed oocysts of Cryptosporidium sp. in the faeces examined by Kinyoun technique: seven children one year old, three, two years old and three, three years old. Among the 23 examined adults, only a 22 years old woman, possibly having an immunocomprometiment, was positive. Clinical and epidemiological aspects were investigated by questionnaires, highlighting the occurrence of the outbreak in a very dry period.O protozoário Cryptosporidium sp. tem sido detectado com freqüência em fezes de crianças com diarréia persistente. Este trabalho teve como objetivo investigar um surto de criptosporidiose, em uma escola de educação infantil, que atende crianças de classe média alta, de 0 a seis anos de idade. O surto foi detectado a partir de amostras fecais não diarréicas encaminhadas ao Instituto Adolfo Lutz através da rede de saúde pública. Das 64 crianças 13 (20,3% apresentaram oocistos de Cryptosporidium nas fezes, examinadas pelo método de Kinyoun, sendo sete crianças com um ano de idade, três com dois anos de idade e três, com três anos de idade. Dos 23 adultos examinados, apenas uma mulher de 22 anos, com possível imunocomprometimento, foi positiva. Aspectos epidemiológicos e clínicos foram investigados por meio de questionários, ressaltando-se que o surto ocorreu em período muito seco.
Robinson, Guy; Chalmers, Rachel M
The use of high resolution molecular tools to study Cryptosporidium parvum and Cryptosporidium hominis intra-species variation is becoming common practice, but there is currently no consensus in the methods used. The most commonly applied tool is partial gp60 gene sequence analysis. However, multi-locus schemes are acknowledged to improve resolution over analysis of a single locus, which neglects potential re-assortment of genes during the sexual phase of the Cryptosporidium life-cycle. Multi-locus markers have been investigated in isolates from a variety of sampling frames, in varying combinations and using different assays and methods of analysis. To identify the most informative markers as candidates for the development of a standardised multi-locus fragment size-based typing (MLFT) scheme to integrate with epidemiological analyses, we examined the published literature. A total of 31 MLFT studies were found, employing 55 markers of which 45 were applied to both C. parvum and C. hominis. Of the studies, 11 had sufficient raw data, from three or more markers, and a sampling frame containing at least 50 samples, for meaningful in-depth analysis using assessment criteria based on the sampling frame, study size, number of markers investigated in each study, marker characteristics (>2 nucleotide repeats) and the combinations of markers generating all possible multi-locus genotypes. Markers investigated differed between C. hominis and C. parvum. When each scheme was analysed for the fewest markers required to identify 95% of all MLFTs, some redundancy was identified in all schemes; an average redundancy of 40% for C. hominis and 27% for C. parvum. Ranking markers, based on the most productive combinations, identified two different sets of potentially most informative candidate markers, one for each species. These will be subjected to technical evaluation including typability (percentage of samples generating a complete multi-locus type) and discriminatory power by
Enemark, Heidi L.; Ahrens, Peter; Bille-Hansen, Vivi
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...
Boyer, Kenneth; Hill, Dolores; Mui, Ernest; Wroblewski, Kristen; Karrison, Theodore; Dubey, J. P.; Sautter, Mari; Noble, A. Gwendolyn; Withers, Shawn; Swisher, Charles; Heydemann, Peter; Hosten, Tiffany; Babiarz, Jane; Lee, Daniel
(See the Editorial Commentary by Linn, on pages 1090–1.) Background. Congenital toxoplasmosis presents as severe, life-altering disease in North America. If mothers of infants with congenital toxoplasmosis could be identified by risks, it would provide strong support for educating pregnant women about risks, to eliminate this disease. Conversely, if not all risks are identifiable, undetectable risks are suggested. A new test detecting antibodies to sporozoites demonstrated that oocysts were the predominant source of Toxoplasma gondii infection in 4 North American epidemics and in mothers of children in the National Collaborative Chicago-based Congenital Toxoplasmosis Study (NCCCTS). This novel test offered the opportunity to determine whether risk factors or demographic characteristics could identify mothers infected with oocysts. Methods. Acutely infected mothers and their congenitally infected infants were evaluated, including in-person interviews concerning risks and evaluation of perinatal maternal serum samples. Results. Fifty-nine (78%) of 76 mothers of congenitally infected infants in NCCCTS had primary infection with oocysts. Only 49% of these mothers identified significant risk factors for sporozoite acquisition. Socioeconomic status, hometown size, maternal clinical presentations, and ethnicity were not reliable predictors. Conclusions. Undetected contamination of food and water by oocysts frequently causes human infections in North America. Risks are often unrecognized by those infected. Demographic characteristics did not identify oocyst infections. Thus, although education programs describing hygienic measures may be beneficial, they will not suffice to prevent the suffering and economic consequences associated with congenital toxoplasmosis. Only a vaccine or implementation of systematic serologic testing of pregnant women and newborns, followed by treatment, will prevent most congenital toxoplasmosis in North America. PMID:22021924
Jenkins, Mark C; Parker, Carolyn; O'Brien, Celia; Persyn, Joseph; Barlow, Darren; Miska, Katarzyna; Fetterer, Raymond
Control of avian coccidiosis is increasingly being achieved by the administration of low doses of Eimeria oocysts to newly hatched chicks. The purpose of this study was to test the efficacy of gel beads containing a mixture of Eimeria acervulina, Eimeria maxima, and Eimeria tenella oocysts as a vaccine to protect broilers raised in contact with litter. Newly hatched chicks were either sprayed with an aqueous suspension of Eimeria oocysts or were allowed to ingest feed containing Eimeria oocysts-incorporated gel beads. Control, 1-day-old chicks were given an equivalent number of Eimeria oocysts (10(3) total) by oral gavage or received no vaccine (nonimmunized controls). All chicks were raised in floor-pen cages in direct contact with litter. At 4 wk of age, all chickens and a control nonimmunized group received a high-dose E. acervulina, E. maxima, and E. tenella challenge infection. Chickens immunized with Eimeria oocysts in gel beads or by spray vaccination displayed significantly (P 0.05) from chickens immunized by oral gavage or from nonimmunized, noninfected controls. Oocyst excretion after Eimeria challenge by all immunized groups was about 10-fold less than in nonimmunized controls. These findings indicate that immunization efficacy of gel beads and spray vaccination is improved by raising immunized chicks in contact with litter.
Codices, Vera; Martins, Catarina; Novo, Carlos; de Sousa, Bruno; Lopes, Ângela; Borrego, Miguel; Matos, Olga
Infection by Cryptosporidium parvum triggers a complex array of innate and adaptive cell mediated immune response, playing an important role in controlling the infection. To date, there are no studies applying the Luminex® xMAP technology to determine profiles of cytokines and immunoglobulins in the context of an infection by C. parvum. In this study, we analyzed these immune mediators in the serum of immunocompetent mice inoculated with C. parvum oocysts, using Luminex, to understand how the immune system responds to an infection by this parasite. Animal sera were also analyzed by ELISA to determine the expressed immunoglobulin isotype profile, and compare the obtained trend with data obtained by Luminex. Specific-pathogen-free BALB/C mice were inoculated with oocysts of C. parvum at days 0 and 22. Peripheral blood was aseptically collected from sacrificed mice on several time points, and immune mediators were evaluated in serum samples. Infection was confirmed by the presence of C. parvum DNA in feces by a nested-PCR assay (60-kDa glycoprotein). Luminex results showed predominance in the secretion of IgG1 and IgG2a, confirmed by ELISA, which also showed that IgG1 is the major immunoglobulin isotype produced during the infection. The analysis of cytokines suggests a preferential Th(1) over the Th(2) response, with increased production of TNF-α, IFN-γ and GM-CSF. This work contributed to a better understanding of the immune response to the infection by C. parvum, as well as demonstrated the advantage of Luminex® xMAP technology to study immune mediators, using small sample volumes. Copyright © 2012 Elsevier Inc. All rights reserved.
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.
Fecal samples were obtained directly from the rectum of each animal. The samples were processed based on the formol ether concentration technique and the saturated sodium chloride flotation method. Cryptosporidium spp. was detected using the modified Ziehl-Neelsen (ZN) method. Significant relationships (p>0.05) ...
Kváč, Martin; Ditrich, Oleg; Kouba, M.; Sak, Bohumil; Vítovec, J.; Květoňová, Dana
Roč. 51, č. 4 (2004), s. 373-374 ISSN 0015-5683 R&D Projects: GA AV ČR IBS6022006 Institutional research plan: CEZ:AV0Z6022909 Keywords : cryptosporidiosis * Cryptosporidium andersoni * experimental infection Subject RIV: DJ - Water Pollution ; Quality Impact factor: 0.837, year: 2004
Full Text Available Considered a zoonosis of utmost importance, cryptosporidiosis has a worldwide distribution and can infect mammals, birds, reptiles, and amphibians. It is caused by a highly resistant protozoan present in the environment and can cause death in immunosuppressed individuals and pups, as well as in farm animals such as cattle and sheep, generating losses. The aim of this study was to investigate the presence of Cryptosporidium spp. in sheep feces from the farms of Western Paraná, which have different management styles, and compare the results with their respective management methods. One hundred and forty-four stool samples were collected (69 from Property 1 and 75 from Property 2 and analyzed using a fecal smear on slides after staining by the modified Ziehl-Neelsen method. Samples tested positive by this method were subjected to nested PCR and the products obtained were sent for sequencing to determine the species. While 82.60% of the samples from Property 1 were tested positive, only 36% of the samples from Property 2 were tested positive. On analyzing the sequencing data, it was observed that the Cryptosporidium species of samples from Property 1 showed high similarity to Cryptosporidium xiaoi and those from Property 2, to Cryptosporidium ubiquitum. The reason for divergence in results can be attributed to differences in management systems adopted by each property, thus showing the importance of detecting carrier animals, as they can contaminate the environment, especially the water sources, and spread the disease to humans and other animals.
Xiao, L.; Ryan, U. M.; Graczyk, T. K.; Limor, J.; Li, L.; Kombert, M.; Junge, R.; Sulaiman, I. M.; Zhou, L.; Arrowood, M. J.; Koudela, Břetislav; Modrý, David; Lal, A. A.
Roč. 70, č. 2 (2004), s. 891-899 ISSN 0099-2240 R&D Projects: GA ČR GA524/00/P015 Institutional research plan: CEZ:AV0Z6022909 Keywords : Cryptosporidium * reptiles * genetic diversity Subject RIV: EG - Zoology Impact factor: 3.810, year: 2004
Objective: To assess the importance of Cryptosporidium parvum and Isospora belli infections as a cause of diarrhoea among patients admitted to the Medical Wards in Queen Elizabeth Central Hospital (QECH) in Blantyre, Malawi. Design: Prospective case control study. Subjects: One hundred and twenty one patients with ...
Traversa, D.; Iorio, R.; Otranto, D.; Modrý, David; Šlapeta, J.
Roč. 22, č. 2 (2008), s. 122-128 ISSN 0890-8508 Institutional research plan: CEZ:AV0Z60220518 Keywords : Cryptosporidium * tortoises * COWP * Testudo * epidemiology * 18S SSU rDNA Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 2.196, year: 2008
Cryptosporidium is said to cause diarrhoea in HIV / AIDS patients. The study was done to determine the prevalence of cryptosporidiosis in Lagos. Stool samples were collected from 193 HIV positive and 200 HIV negative (control) patients presenting with diarrhoea at LUTH. The patient or a close relative filled a ...
Mølbak, Kare; Andersen, M; Aaby, Peter
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...
Medema GJ; Theunissen J; MGB
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
Wastewater reuse is becoming increasingly important for supplementing drinking water supply needs and/or to reduce costs in many communities around the world. However, wastewater reuse can result in a potential transmission route for infectious agents. Therefore, the occurrence of Cryptosporidium was assessed in a ...
Full Text Available Genetic evidence for the occurrence of two Cryptosporidium parvum subgroups is presented. This evidence is based on restriction fragment length polymorphism analysis of several independent loci. Sequence analysis of the b -tubulin intron revealed additional polymorphism. The stability of the genetic profiles following passage of C. parvum isolates between different hosts was investigated.
Enemark, Heidi L.; Bille-Hansen, Vivi; Lind, Peter
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...
Kváč, Martin; Němejc, K.; Kestřánová, M.; Květoňová, Dana; Wagnerová, Pavla; Kotková, Michaela; Rost, M.; Samková, E.; McEvoy, J.; Sak, Bohumil
Roč. 202, 3-4 (2014), s. 330-334 ISSN 0304-4017 R&D Projects: GA MŠk(CZ) LH11061 Institutional support: RVO:60077344 Keywords : Cryptosporidium scrofarum * molecular analyses * transmission studies * susceptibility * infection * pigs Subject RIV: EE - Microbiology, Virology Impact factor: 2.460, year: 2014
Diseases in particular parasitic infestation is among the drawbacks to profitable pig production since parasites compromise the production and reproduction performance of infested pigs. The objective of this study was to estimate the prevalence and associated risk factors for Ascaris and Cryptosporidium infestations in pigs ...
Schets FM; Leenen EJTM; MGB
In vivo en in vitro modellen voor kweek van Cryptosporidium geven informatie over de infectiviteit van oocysten in ruw water en drinkwater en kunnen de identificatie van middelen met een anticryptosporidiele werking vereenvoudigen. In vivo modellen zijn echter duur, onpractisch en niet in elk
A single faecal sample per animal was collected from pigs, goats, dogs, ducks, chickens and pigeons and analysed by Merifluor Cryptosporidium/Giardia immunofluorescence antibody assay for the simultaneous detection of these parasites. The faecal consistency was noted and scored as non-diarrhoeic or diarrhoeic.
Cryptosporidiosis is an important zoonotic disease caused by the protozoa Cryptosporidium. Infections in birds are mainly caused by three species C. meleagridis, C. baileyi, and C. galli. C. meleagridis is the third most common cause of cryptosporidiosis in immunocompromised and immunocompetent huma...
Medema, Gerriet Jan
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