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Development of Quantification and Genotyping methods for Cryptosporidium in water by Quenching Probe PCR followed by RFLP
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- MASAGO Yoshifumi
- Department of Urban Engineering, Graduate School of Engineering, The University of Tokyo
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- OGUMA Kumiko
- Department of Urban Engineering, Graduate School of Engineering, The University of Tokyo
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- KATAYAMA Hiroyuki
- Department of Urban Engineering, Graduate School of Engineering, The University of Tokyo
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- OHGAKI Shinichiro
- Department of Urban Engineering, Graduate School of Engineering, The University of Tokyo
Bibliographic Information
- Other Title
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- 消光型蛍光プローブを用いたリアルタイムＰＣＲ法による水中のクリプトスポリジウムの定量および種別判定手法の開発
Description
A new method was developed to quantify Cryptosporidium in water. Quenching Probe PCR (QProbe-PCR) technique could successfully amplify approximately 1280bp of Cryptosporidium 18S rDNA from a sample with as low as 60 [oocysts/tube] of Cryptosporidium parvum bovine genotype. QProbe-PCR showed high accuracy and high sensitivity compared to Real Time PCR with TaqMan probe.<BR>QProbe-PCRh as an advantaget hat the PCR products can be applied for molecular characterization. Arestriction fragment length polymorphism (RFLP) technique was used to distinguish Cryptosporidium species and genotypes. Five species (C. parvum bovine genotype, C. parvum human genotype, C. meleagridis, C. felis and C. muris) could be distinguished by the RFLP with restriction enzymes Ssp I, Vsp I and Sty I. The Sty I successfully differentiated C. muris calf genotype (also known as C. andersoni) and C. muris mouse genotype. Database-based analysis revealed that 8 species out of 10 could be distinguished by RFLP with these three restriction enzymes.<BR>QProbe-PCR-RFLP techniques can provide information on the genotype as well as the quantity of Cryptosporidium from the same sample. This technique can be a useful tool for waterborne risk assessment of Cryptosporidiosis.
Journal
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- ENVIRONMENTAL ENGINEERING RESEARCH
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ENVIRONMENTAL ENGINEERING RESEARCH 41 311-319, 2004
Japan Society of Civil Engineers
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Keywords
Details 詳細情報について
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- CRID
- 1390282679089503232
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- NII Article ID
- 130003949381
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- ISSN
- 1884829X
- 13415115
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- Text Lang
- ja
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- Data Source
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- JaLC
- CiNii Articles
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- Abstract License Flag
- Disallowed
