Evaluation of TaqMan real-time PCR for the detection of viable Cryptosporidium parvum oocysts in environmental water samples
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Detection and enumeration of Cryptosporidium oocysts in environmental water samples by Real-time PCR assay
Introduction: The protozoan parasite, Cryptosporidium Spp., widely spreads in both raw and drinking waters. It is the causative agents of waterborne diarrhea and gastroenteritis in the world. In the present study, a molecular assay was used for the detection and quantification of Cryptosporidium oocysts in environmental water samples. Materials and methods: Thirty surface water samples wer...
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We recently described a reverse transcription-PCR (RT-PCR) for detecting low numbers of viable Cryptosporidium parvum oocysts spiked into clarified environmental water concentrates. We have now modified the assay for direct analysis of primary sample concentrates with simultaneous detection of viable C. parvum oocysts, Giardia cysts, and a novel type of internal positive control (IPC). The IPC ...
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Cryptosporidium parvum is a common intestinal protozoan parasite infecting humans and a wide range of animals, whose diagnostics present considerable difficulties. These arise from the exceptionally robust nature of the oocyst’s walls, which necessitates more stringent treatments for disruption and recovery of DNA for analysis using molecular methods. In the case of water, which is the major so...
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Extraction of high-quality mRNA from Cryptosporidium parvum is a key step in PCR detection of viable oocysts in environmental samples. Current methods for monitoring oocysts are limited to water samples; therefore, the goal of this study was to develop a rapid and sensitive procedure for Cryptosporidium detection in soil samples. The efficiencies of five RNA extraction methods were compared (mR...
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A rapid detection method that is both quantitative and specific for the water-borne human parasite Cryptosporidium parvum is reported. Real-time polymerase chain reaction (PCR) combined with fluorescent TaqMan technology was used to develop this sensitive and accurate assay. The selected primer-probe set identified a 138-bp section specific to a C. parvum genomic DNA sequence. The method was op...
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