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Combined immunomagnetic separation-molecular beacon-reverse transcription-PCR assay for detection of hepatitis A virus from environmental samples.

ABSTRACT: In this study, a molecular-beacon-based real-time reverse transcription (RT)-PCR assay was developed to detect the presence of hepatitis A virus (HAV) in environmental samples. A 125-bp, highly conserved 5' noncoding region of HAV was targeted. The sensitivity of the real-time RT-PCR assay was tested with 10-fold dilutions of viral RNA, and a detection limit of 1 PFU was obtained. The specificity of the assay was demonstrated by testing with other environmental pathogens and indicator microorganisms, and only HAV was positively identified. When combined with immunomagnetic separation, the real-time RT-PCR assay successfully detected as few as 20 PFU in seeded groundwater samples. Because of its simplicity and specificity, this assay has broad applications for the rapid detection of HAV in contaminated foods or water.

SUBMITTER: Abd El Galil KH 

PROVIDER: S-EPMC444796 | biostudies-literature | 2004 Jul

REPOSITORIES: biostudies-literature

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Combined immunomagnetic separation-molecular beacon-reverse transcription-PCR assay for detection of hepatitis A virus from environmental samples.

Abd El Galil Khaled H KH   El Sokkary M A MA   Kheira S M SM   Salazar Andre M AM   Yates Marylynn V MV   Chen Wilfred W   Mulchandani Ashok A  

Applied and environmental microbiology 20040701 7

In this study, a molecular-beacon-based real-time reverse transcription (RT)-PCR assay was developed to detect the presence of hepatitis A virus (HAV) in environmental samples. A 125-bp, highly conserved 5' noncoding region of HAV was targeted. The sensitivity of the real-time RT-PCR assay was tested with 10-fold dilutions of viral RNA, and a detection limit of 1 PFU was obtained. The specificity of the assay was demonstrated by testing with other environmental pathogens and indicator microorgan  ...[more]

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