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Aquatic Microbial Ecology

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AME 47:209-212 (2007)  -  doi:10.3354/ame047209

Tungstate compounds as an alternative to uranyl acetate for enumerating viral-infected prokaryote cells in aquatic systems

A. S. Pradeep Ram1,3, T. Nagata1,*, Y. Tomaru2, K. Nagasaki2

1Centre for Ecological Research, Kyoto University, 2-509-3 Hirano, Otsu, Shiga 520-2113, Japan
2Harmful Algal Bloom Division, National Research Institute of Fisheries and Environment of Inland Sea, Fisheries Research Agency, 2-17-5 Maruishi, Hatsukaichi, Hiroshima 739-0452, Japan
3Present address: Laboratoire de Biologie des Protistes, Université Blaise Pascal (Clermont-Ferrand II), UMR CNRS 6023, 63177 Aubiere cedex, France
*Corresponding author. Email:

ABSTRACT: We examined whether tungstate compounds (sodium phosphotungstate and sodium silicotungstate) can be used as a safe alternative to uranyl acetate (UA) for the purpose of detecting and counting viruses inside aquatic prokaryotic cells using transmission electron microscopy (TEM). Tungstate compounds used as negative stains at neutral to slightly acidic pH are less effective in the detection of viral particles inside the cells. However, the use of tungstate compounds at low pH (3 or 4) substantially improved viral images, being positively contrasted inside the cells. With this improvement, the estimates of the frequency of viral-infected cells derived by the tungstate method agreed well with those determined by the use of UA for both lake and marine waters. We also found that the average burst size of the lake sample differed little between tungstate and the UA approach; however, the average burst size in a marine sample was underestimated by the tungstate method because of the failure to resolve highly packed viral particles. The above results suggest that the use of tungstate compounds at low pH can be a useful alternative to the UA approach for estimating the frequency of viral-infected cells in aquatic samples, but results on the burst size need to be interpreted cautiously when the burst size is high.

KEY WORDS: Tungstate compounds · Uranyl acetate · Transmission electron microscopy · Frequency of viral infected cells · Burst size · Prokaryotes · Viruses

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