A Method for the Detection of Cryptosporidium parvum Oocysts in Milk Based on Microfiltration and Real-Time Polymerase Chain Reaction

A method for the detection of Cryptosporidium parvum oocysts in milk was developed on the basis of optimizing microfiltration and elution of the material from the filter, and using a previously developed highly sensitive downstream detection by real-time polymerase chain reaction. The method involve...

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Bibliographic Details
Published in:Food analytical methods 2011-03, Vol.4 (1), p.116-120
Main Authors: Minarovičová, Jana, Lopašovská, Janka, Valík, Ľubomír, Kuchta, Tomáš
Format: Article
Language:English
Subjects:
Analytical Chemistry
Chemistry
Chemistry and Materials Science
Chemistry/Food Science
Food Science
Microbiology
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Summary:A method for the detection of Cryptosporidium parvum oocysts in milk was developed on the basis of optimizing microfiltration and elution of the material from the filter, and using a previously developed highly sensitive downstream detection by real-time polymerase chain reaction. The method involves heating of milk to 40°C, microfiltration through a membrane microfilter made of a mixture of cellulose acetate and cellulose nitrate (pore size, 3.0 μm), elution of the material from the filter by a solution containing sodium pyrophosphate and Tween 80 in a shaker, rapid DNA extraction using a Chelex-based agent, and single-tube nested real-time PCR. The detection limit of the method is 10 C. parvum oocysts per 100 ml of milk. The developed method may be useful for specific and sensitive control of contamination of milk by C. parvum oocysts.
ISSN:1936-9751
1936-976X
DOI:10.1007/s12161-010-9141-9