AEM
Home Help [Feedback] [For Subscribers] [Archive] [Search] [Contents]
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Shieh, Y.-S. C.
Right arrow Articles by Baric, R. S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Shieh, Y.-S. C.
Right arrow Articles by Baric, R. S.
Agricola
Right arrow Articles by Shieh, Y.-S. C.
Right arrow Articles by Baric, R. S.

Next Article 

Applied and Environmental Microbiology, November 1999, p. 4709-4714, Vol. 65, No. 11
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

A Method To Detect Low Levels of Enteric Viruses in Contaminated Oysters

Y.-S. Carol Shieh,1,* Kevin R. Calci,1 and Ralph S. Baric2

Gulf Coast Seafood Laboratory, U.S. Food and Drug Administration, Dauphin Island, Alabama 36528,1 and Program in Infectious Diseases, Department of Epidemiology, University of North Carolina, Chapel Hill, North Carolina 275992

Received 15 April 1999/Accepted 4 August 1999

Direct isolation and identification of pathogenic viruses from oysters implicated in gastroenteritis outbreaks are hampered by inefficient methods for recovering viruses, naturally occurring PCR inhibitors, and low levels of virus contamination. In this study we focused on developing rapid and efficient oyster-processing procedures that can be used for sensitive PCR detection of viruses in raw oysters. Poliovirus type 3 (PV3) Sabin strain was used to evaluate the efficacy of virus recovery and the removal of PCR inhibitors during oyster-processing procedures. These procedures included elution, polyethylene glycol precipitation, solvent extraction, and RNA extraction. Acid adsorption-elution in which glycine buffer (pH 7.5) was used was found to retain fewer inhibitors than direct elution in which glycine buffer (pH 9.5) was used. RNA extraction in which a silica gel membrane was used was more effective than single-step RNA precipitation for removing additional nonspecific PCR inhibitors. The final 10-µl volume of RNA concentrates obtained from 2 g of oyster tissue (concentration factor, 200-fold) was satisfactory for efficient reverse transcription-PCR detection of virus. The overall detection sensitivity of our method was 1 PFU/g of oyster tissue initially seeded with PV3. The method was utilized to investigate a 1998 gastroenteritis outbreak in California in which contaminated oysters were the suspected disease transmission vehicle. A genogroup II Norwalk-like virus was found in two of three recalled oyster samples linked by tags to the harvest dates and areas associated with the majority of cases. The method described here improves the response to outbreaks and can be used for rapid and sensitive detection of viral agents in outbreak-implicated oysters.

* Corresponding author. Mailing address: Gulf Coast Seafood Laboratory, U.S. Food and Drug Administration, P.O. Box 158, Dauphin Island, AL 36528. Phone: (334) 690-3407. Fax: (334) 694-4477. E-mail: ycs{at}vm.cfsan.fda.gov.

Applied and Environmental Microbiology, November 1999, p. 4709-4714, Vol. 65, No. 11
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:



Home Help [Feedback] [For Subscribers] [Archive] [Search] [Contents]
J. Bacteriol. Microbiol. Mol. Biol. Rev. Eukaryot. Cell All ASM Journals

Copyright © 1999 by the American Society for Microbiology. All rights reserved.