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Applied and Environmental Microbiology, July 2006, p. 4767-4774, Vol. 72, No. 7
0099-2240/06/$08.00+0     doi:10.1128/AEM.00297-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Assessment of Factors Influencing Direct Enumeration of Viruses within Estuarine Sediments

Rebekah R. Helton,1 Ling Liu,2 and K. Eric Wommack1,3*

Department of Plant and Soil Sciences, Delaware Biotechnology Institute, University of Delaware, Newark, Delaware 19711,1 Department of Food and Resource Economics, University of Delaware, Newark, Delaware 19711,2 College of Marine Studies, University of Delaware, Lewes, Delaware 199583

Received 6 February 2006/ Accepted 5 May 2006

Accurate enumeration of viruses within environmental samples is critical for investigations of the ecological role of viruses and viral infection within microbial communities. This report evaluates differences in viral and bacterial direct counts between estuarine sediment samples which were either immediately processed onboard ship or frozen at –20°C and later processed. Viral and bacterial abundances were recorded at three stations spanning the length of the Chesapeake Bay in April and June 2003 within three sediment fractions: pore water (PW), whole sediment (WS), and sediment after pore water removal (AP). No significant difference in viral abundance was apparent between extracts from fresh or frozen sediments. In contrast, bacterial abundance was significantly lower in the samples subjected to freezing. Both bacterial and viral abundance showed significant differences between sediment fractions (PW, WS, or AP) regardless of the fresh or frozen status. Although pore water viral abundance has been used in the past as a measurement of viral abundance in sediments, this fraction accounted for only ca. 5% of the total sediment viral abundance across all samples. The effect of refrigerated storage of sediment viral extracts was also examined and showed that, within the first 2 h, viral abundance decreased ca. 30% in formalin-fixed extracts and 66% in unfixed extracts. Finally, the reliability of direct viral enumeration via epifluorescence microscopy was tested by using DNase treatment of WS extractions. These tests indicated that a large fraction (>86%) of the small SYBR gold fluorescing particles are likely viruses.

* Corresponding author. Mailing address: Delaware Biotechnology Institute, 15 Innovation Way, Rm. 148, Newark, DE 19711. Phone: (302) 831-4362. Fax: (302) 831-3447. E-mail: wommack{at}dbi.udel.edu.

Applied and Environmental Microbiology, July 2006, p. 4767-4774, Vol. 72, No. 7
0099-2240/06/$08.00+0     doi:10.1128/AEM.00297-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Helton, R. R., Wommack, K. E. (2009). Seasonal Dynamics and Metagenomic Characterization of Estuarine Viriobenthos Assemblages by Randomly Amplified Polymorphic DNA PCR. Appl. Environ. Microbiol. 75: 2259-2265 [Abstract] [Full Text]  


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