Full Description

Field testing of a strategy using immunomagnetic separation (IMS) of naturally occurring C. parvum oocysts from environmental water samples followed by infectivity determination using integrated cell culture-PCR (CC-PCR) was initiated. Over 100 each raw source water and filter backwash water grab samples from twenty-five sites in the U.S. have been analyzed. Oocyst seeded raw and filter backwash water samples were also used to evaluate recovery efficiencies and performance of the CC-PCR protocol with different water quality matrices. The grab samples were concentrated by centrifugation, concentrates split and purified by IMS. An acidified Hank's balanced salt solution (HBSS) containing 1% trypsin was used for the dissociation of captured oocysts from the IMS beads since the manufacturer recommended 0.1 N HC1 dissociation may adversely affect oocyst viability. In vitro HCT-8 cell culture of IMS purified oocysts was performed in 96-well microtiter plates and infected cells were detected using PCR primers specific for C. parvum. CC-PCR has detected infectious Cryptosporidium parvum in 6 raw and 9 filter backwash water samples. All CC-PCR positive samples were confirmed by cloning and DNA sequence analysis of the PCR products.

Product Details

Published: 01/01/1998 ISBN(s): 0898679796 Number of Pages: 6 File Size: 1 file , 38 KB