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A study conducted to better estimate the annual rate of environmental loading of Cryptosporidium spp. by colonies of California ground squirrels, to better map the distribution of genotypes for the host species is described. Sufficient numbers of oocysts were not available to conduct a mouse infectivity trial, as Sbey03a is a rare type of genotypes in the sample.

The five member filters consisting of Pall Life Sciences Envirochek (EC) standard filtration, Envirochek high-volume (EC-HV) membrane filters, the Millipore flatbed membrane filter, Sartorius flatbed membrane filter (SMF) and Filta-Max (FM) depth filter were evaluated for studying their effectiveness in the recovery of Crytosporidium parvum oocysts and Giardia lamblia cysts. Results indicate that the Giardia lamblia cysts were recovered at a higher rate as compared to the Crytosporidium parvum oocysts.

Cryptosporidium Oocysts that are present in animal fecal deposits are linked to increases in pathogen concentration in streams and reservoirs. Dispersion and transportation of Cryptosporidium oocysts from fecal pats were studied under simulated rainfall conditions and results are discussed and tabulated.

The genetic diversity of cryptosporidium in reptiles was analyzed using PCR-restriction fragment length polymorphism and sequence analysis of a small subunit rRNA gene. In the study samples of snakes, lizards and tortoises were analyzed and results suggest extensive genetic diversity.

Results demonstrate that about 32.2% of the oocyst-positive raw and 41% of oocyst-positive final water samples do not contain 4',6'-Diamidino-2-phenylindole-positive nuclei. This method based on the presence of one or more fluorescent nuclei in a positive sample provides an enhanced fluorogenic detection method.

Many ducks carry Cryptosporidium and Giardia species but not C. parvum or G. lamblia. Of 69 fecal samples taken from wild ducks in New Mexico, 49% tested positive for Cryptosporidium and 28% tested positive for Giardia. C. parvum and G. lamblia are the species most often associated with illness in humans.

Increased water potential of soil can increase the rate of degradation of Cryptosporidium parvum oocysts at all temperatures. This is important because many animals infected with the parasite may excrete the oocysts on soil in watersheds that supply drinking water.

Several sample preparation methods for extracting high-quality DNA for PCR were evaluated. For Escherichia coli, Xtra Amp tube, Isocode paper, Instagene matrix, and PrepMan matrix methods were all suitable. For Cryptosporidium parvum, Xtra Amp and Instagene methods were suitable.

Capturing Cryptosporidium parvum oocysts in water using the cellulose acetate membrane dissolution filtration system can render the oocysts non-infective. This would make it difficult for researchers to determine the viability of the oocysts.

Monchromatic, low-pressure UV radiation can inactivate Cryptosporidium oocysts and the oocysts do not seem to be capable of repairing the CNA damage. Cryptosporidium is resistant to other forms of disinfection, including chemical disinfection.

Some bacterial spores are more sensitive to chlorine dioxide than Cryptosporidium parvum and therefore should not be used as indicator organisms. In addition, oocyst purification methods may differ, as purified oocysts from three different supplies had different sensitivities to chlorine dioxide.

Some California ground squirrels shed over 50,000 Cryptosporidium parvum oocysts per gram of feces. Males shed greater numbers of oocysts than females, which could facilitate the dissemination of the organism because males are more likely to leave the family colony.

Researchers report the first known use of MALDI-TOF mass spectroscopy to analyze protozoal oocysts. They found reproducible patterns of spectral markers that could distinguish Cryptosporidium parvum from other Cryptosporidium species.

Research has been conducted on the heat shock protein genes. The characterazation of these genes' nucleotide sequences has been carried out.

Research has been conducted on the Crystosporidium isolates' genetic and phylogenetic characterization. Results indicate that dog-derived Crystosporidium isolates have a distinct genotype.

Results show that Cryptosporidium meleagridis is distinct from others in the family of coccidian parasites and is widely distributed with a broad host range. Data point out that birds are susceptible for these parasite.

Research was conducted to quantify infectious Cryptosporidium parvum oocysts using the most-probable-number (MPN) method of enumerating microorganisms. The approach involved creating a dilution series, incubating the samples for an adequate period, plating in replicate and scoring the presence of growth. Results provide evidence that the focus detection method-MPN approach is an excellent and reproducible assay for quantifying oocyst infectivity in vitro.

A direct immunofluorescence assay (DFA) was contrasted with an immunomagnetic separation (IMS) assay combined with immunofluorescent microscopy for their ability to detect low concentrations of Cryptosporidium parvum oocysts in adult bovine feces. IMS-DFA resulted in a 2-log-unit increase in sensitivity compared to DFA alone. The higher sensitivity of the IMS-DFA resulted from testing 2 g of fecal material instead of the 13 to 19 mg of fecal material tested in the DFA. The increased sensitivity was not attributable to a higher percent recovery.

A two-year study of the survival kinetics of oocysts in field settings on three watershed farms in New York, NY, is reported. Wastes from neonatal calves in these farms were being managed as part of dairy operations. A small-volume sentinel system was used to evaluate the effects of environmental stresses on survival of Cryptosporidium parvum oocysts in soil and animal wastes. Results suggested that manure piles containing C paevum oocysts can hasten inactivation and thus reduce the numbers of infective oocysts before wastes are spread in the soil environment.

A new infection caused by Cryptosporidium felis, aside from known infections with C muris and C parvum, was identfiied in a cow bred on the Bystra farm in northern Poland. The identfication was made on the basis of the size of the oocysts and analysis of the molecular sequence of the variable region of the small-subunit rRNA. This finding provides important insight into the complex host specificity and circulation in the environment of Cryptosporidium species.

Evidence from biological characteristics and host specificity studies show that species differentiation occurs in Cryptosporidium parasites. This observation was further supported by a phylogenetic analysis of the parasite genus Cryptosporidium wherein the small-subunit rRNA genes of Cryptosporidium parvum, Cryptosporidium muris, Cryptosporidium baileyi and Cryptosporidium serpentis were characterized. The Cryptosporidium species produced two clades, with Cryptosporidium baileyi and Cryptosporidium parvum belonging to one clade and Cryptosporidium muris and Cryptosporidium serpentis belonging to the other.

The viability of waterborne Cryptosporidium parvum oocysts can be assessed using beta-tubulin mRNA as marker. To prove this observation, rRNA and mRNA were investigated in a reverse transcription-PCR assay as potential markers of oocyst viability. mRNA levels decreased gradually after prolonged incubation of initially viable oocysts at room temperature, which coincided with the loss of oocyst infectivity to neonatal mice under study. The postmortem decay of two mRNA species shows that reverse transcription-PCR analysis can elucidate the viability of Cryptosporidium parvum oocysts.

Research was conducted to evaluate the effect of immunomagnetic separation (IMS) on oocyst infectivity by means of in vitro cell culture assay combined with mRNA extraction and reverse transcriptase PCR. Two IMS kits for Cryptosporidium parvum were compared for recovery of oocysts from environmental samples. Results provide evidence that oocysts that were recovered by IMS retained their infectivity.

Research was conducted to examine the effects of sample turbidity on the recovery of Cryptosporidium parvum oocysts using two immunomagnetic separation (IMS) kits. The objectives of the study was to identify an IMS kit that generated high purified oocyst suspensions, to determine turbidity levels and to use the IMS kit for the evaluation of the effects of age on the recovery of C. parvum oocysts. Results indicate that IMS is a promising alternative to flotation procedures for the recovery of oocysts from turbid water matrices.

Research was conducted to examine the sedimentation velocities of Cryptosporidium parvum oocysts and Giardia lamblia cysts using measurements of density, (oo)cyst size, and viscosity of the sedimentation medium. The objective was to observe whether the Stokes' law can describe the sedimentation kinetics of freely suspended cysts of G. lamblia and oocysts of C. parvum. Results indicate that particle attachment has an influence on the behavior of (oo)cysts in drinking water treatment processes and on their sedimentation in surface water.

Research was conducted to examine the effect of pretreatment and experimental conditions on the hydrophobicity and electrophoretic mobility of Cryptosporidium parvum oocysts. A further objective was to measure the effects of solution ionic strength on the hydrophobicity of polystyrene beads. Results indicate that oocysts purified using the EAPS method displayed an electrophoretic mobility that strongly depended on pH.

A simplified version of the randomly amplified polymorphic DNA technique was used to genetically characterize Cryptosporidium parvum. This approach involved the introduction of a genomic DNA isolation method that used in vitro excystation and specific immunomagnetic separation of Cryptosporidium parvum oocysts. Results indicated that it was effective in isolating an estuary C. parvum isolate from several bovine isolates.

Oysters in natural waters harbor infectious Cryptosporidium parvum oocysts. This was demonstrated in a study of the survival of oocysts of Cryptosporidium parvum in artificial seawater of various salinities and various temperatures. Those placed in artificial seawater at salinities of 10, 20 and 30 ppt at 10 degrees Centigrade and at 10 ppt at 20 degrees Centigrade were infectious after 12 weeks. Oocysts placed in seawater at 20 ppt and 30 ppt at 20 degrees Centigrade were infectious for eight and four weeks, respectively.

The natural production and release of free ammonia may have an impact on the viability of Cryptosporidium parvum oocysts in soil and water microhabitants. A study was conducted to determine how the survival of oocysts is affected by increased free ammonia concentrations and times of exposure. The findings indicate that the inactivation of oocysts in ammonia-containing environments may be increased by environmentally relevant concentrations of free ammonia.

A study was conducted on Asian freshwater clams exposed to water contaminated with infectious Cryptosporidium parvum oocysts. No oocysts were detected in the water 24 hours 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. Positive samples were obtained when gills and gastrointestinal tracts were processed with immunofluorescent antibody. Clam-released oocysts were surrounded by feces, no free oocysts or oocysts dissociated from fecal matter were observed.

A cross-sectional study was conducted on the risk factors and prevalence of Giardia sp cysts and Cryptosporidium parvum oocsyts in populations of feral pigs in western California. Results show that, of the 221 feral pigs studied, 12 were shedding Cryptosporidium oocysts and 17 were shedding Giardia sp cysts. Results also indicate that feral pigs may cause surface water protozoal contamination.

The capacity of dye permeability assay to identify Cryptosporidium parvum oocysts was examined by comparing the test to an in vitro excystation assay. The assays were used on sodium hypochlorite-permeabilized formaldehyde-fixed and untreated oocysts. Results show that dye permeability viability testing is not based on metabolic activity potential but on oocyst wall permeability. Results also verify the use of dye permeability assay in predicting oocyst infectivity.

An alternative disinfection system composed of an electrochemically produced mixed-oxidant solution was examined for inactivation of both Cryptosporidium parvum oocysts and Clostridium perfringens spores. The disinfection efficacy of the mixed-oxidant solution was compared to that of free chlorine. Findings showed that the mixed-oxidant solution was considerably more effective than free chlorine in inactivating both microorganisms. Free chlorine produced no quantifiable inactivation of C. parvum oocysts and C. perfringens spores.

Cryptosporidium parvum oocysts obtained from the feces of Peking ducks, innoculated with the oocysts, produces severe infection in neonatal mice. Histological sections of the ducks fail to reveal life-cycle stages of the cryptosporidia. The number of oocyst shells in the feces is less than that of intact oocysts. The number of intact oocyst shells decreases from day two to day five post inoculation. This indicates that the viability and infectivity of the oocysts is unaffected by passage through the duck's intestine.

Raw water samples from the black mountain irrigation district, (BMID), and the vernon irrigation district, (VID), contain Giardia cysts. The parasite contamination is higher for the BMID watershed than the VID because cattle can access the creeks in the former. The peak concentration of both parasites coincides with calving activity. The downstream creek of cattle ranch has more Giardia cysts and Cryptosporidium oocysts than the upstream creek. The fecal samples from the two geographically adjacent areas are Giardia positive but Cryptosporidium negative.

The low adhesion of oocysts of the parasite, Cryptosporidium parvum, to octane indicates that Cryptosporidium sp. have low hydrophobic properties. Analysis of hydrophobicity and zeta values indicates that pH and ionic strength affect the hydrophobicity. The maximum adhesion of the oocysts to octane is only 40%. The zeta potentials are pH-dependent and become less negative as pH drops.

The Cryptosporidium DNA recovery-PCR detection method effectively identifies the genomic DNA of the pathogenic protozoa Cryptosporidium in bovine feces. The test is more sensitive than the commercially used Color Vue-Cryptosporidium EIA test. The DNA is extracted by freezing and thawing dilute feces solutions followed by incubation and centrifugation. The PCR amplification involves the hybridization of the DNA with an internal oligonucleotide probe. No harmful wastes are produced during the procedure.

A polymerase chain reaction (PCR) protocol, that involves hybridization with a chemiluminescent probe, detects the oocysts of the parasite Cryptosporidium in environmental samples such as wastewater, surface water, and drinking water. The method is faster and more reliable than the generally used immunofluorescence method. Specimens are prepared for the PCR assay by pretreatment followed by a freeze-thaw procedure. Unlike the immunofluorescence assay, result of this PCR assay is not influenced by empty oocysts.

Some species of algae interfere with tests for the protozoan parasites Giardia and Cryptosporidium by emitting background fluorescence, but goat serum may eliminate this interference when it is added to the test mixture. Researchers tested 54 species of algae for interference with a fluorescence test for the parasites and found that 24 species emitted fluorescence. Goat serum prevented fluorescent emission from most of these species but did not affect the fluorescence of Giardia cysts and Cryptosporidium eggs.

Propidiumiodide (PI) and 4',6-diamidino-2-phenylindole (DAPI) are fluorogenic vital dyeswhich were previously shown to be promising candidates for assessing viability of protozoan cysts and oocysts. The inclusion or exclusion of these dyes were compared to in vitro excystation of Cryptosporidium parvum oocysts. The resultsshowed that oocysts which are permeable to DAPI but not to PI will excyst usinga standard excystation protocol. Simultaneous staining using DAPI and PI is a useful protocol to determine the viability of C. parvum oocysts.

PCR-restriction fragment length polymorphism analysis may not be able to distinguish some genotypes of Cryptosporidium parvum in a sample that contains several different genotypes. This was the conclusion of researchers who used the technique to analyze combinations of 10 C. parvum oocysts containing various ratios of genotypes I and II.

A high-rate algal pond can reduce the infectivity of Cryptosporidium parvum oocysts by 97%. These ponds are used to treat wastewater and consist of algae to provide oxygen for biodegradation and bacteria to provide nutrients for the algae.

The method detection limits (MDL) of polymerase chain reaction (PCR) and immunofluorescence assay (IFA) for Cryptosporidium parvum oocysts in soil were investigated. Experiments involved soil characterization and seeding, recovery of oocysts from soils, DNA extraction and purification and PCR amplification. Results showed that the recovery efficiency for IFA is stable across orders of magnitude of seeding levels. In contrast, PCR is not effective as a screening step for soil studies of oocyst transport.

Research was conducted to determine if nucleic acid amplification with primers specific for Cryptosporidium parvum followed by automated DNA sequence analysis of the amplicons could discriminate C parvum isolates derived from different mammalian sources throughout California. A previously set developed set of PCR primers was used for the experiments. Results indicate that there was no sufficient polymorphism within the amplicon to allow for reliable differentiation between C parvum isolates obtained within California.

A study on Cryptosporidium sp. infections in green turtles or Chelonia mydas was conducted to determine the sources of marine waterborne oocysts in the Hawaiian Islands. The study identified Cryptosporidium sp. oocysts in fecal and intestinal samples from the turtles. Results show that positive reactions were present and that the range of oocyst concentrations was similar to those reported from snakes.

Studies done to develop a specific, sensitive and photostable detection method for C.parvum oocysts by using luminescence QD-streptavidin together with biotinylated anti-Cryptosporidium sp.monoclonal antibody (MAb) are reported. It is demonstrated that the QD-streptavidin with biotinylated MAb was able to provide a sensitive technique for detecting C.parvum oocysts with the advantage of having a high photobleaching threshold.

The chamelea gallina clams that were accumulated from the mouth of the rivers beside the Adriatic Sea, which is located in the centre of Italy, were identified to anchor the cryptosporidium parvum, which serially forms a part of the zoonotic program. Cryptosporidium is segregated as a powerful tool since it helps to scrutinize the patterns of the program and the epidemiology of these microorganisms.

Thirteen different locations were checked in Ohio and Illinois for the free-living waterfowl Canada geese due to the existence and circulation of cryptosporidum spp. in the fecal droppings of the above geese. The outcome of this check, which was conducted, is that the Canada geese could only by chance transmit the cryptosporidia infection to human beings and thus do not play a significant part in the process of transmitting the pathogen from animals to humans.

The U.S. Environmental Protection Agency method 1623 is largely utilized to check the water sources and to check the supply of drinking water for cryptosporidium ooycsts. The revival between the two seeding processes was very much interconnected and not so distinctly different, but the revival with the help of altered oocysts was as good as the revival by traditional seeding method.

Genetic difference was identified among the Cryptosporidium species from rural and urban isolates in New Zealand, the Cryptosporidium hominis was more common in the urban area and the Cryptosporidium parvum bovine genes were more common among the rural areas. It was further noticed that the transmission cycles also changed like the anthroponotic cycle in autumn and the zoonotic cycle in spring.