Name: Ashley Whelpley

Email: awhelple@uci.edu

Author: Ashley L. Whelpley*, Jessica L. Clasen, Jennifer B.H. Martiny

Author affiliation: Department of Ecology and Evolutionary Biology, University of California Irvine, Irvine, CA, USA * Presenting author

Abstract title: A Comparison of Culture-Dependent and Culture-Independent Techniques for Studying the Genetic Diversity of Cyanomyoviridae

Absstract:

The genetic diversity of cyanophages has been studied by culture-dependent and culture-independent techniques. Both techniques have inherent biases; culture-dependent techniques select for viruses that infect a particular host, while PCR-based culture-independent techniques select for viruses that are targeted by particular primers. Here, we directly compare the two techniques to investigate the relative extent of cyanomyovirus diversity that each captures. We collected seawater from Crystal Cove State Beach, CA on two sampling dates (May and October 2008). For the culture-dependent assay, we isolated viruses on Synechococcus WH7803 using dilution to extinction, followed by plaque purification. Cyanomyovirus diversity and composition was then determined by PCR-amplification of the g20 gene, which encodes for a protein involved in capsid assembly. For the culture-independent technique, we concentrated environmental virioplankton on an Anotop filter; we then PCR amplified the g20 gene, constructed a clone library, and sequenced ~96 clones. The diversity revealed by the culture-dependent technique was restricted phylogenetically; most (94%) of the isolates fell within one cluster. In contrast, a majority of sequences from the culture-independent technique fall outside this isolate cluster (>80% of May sequences and >95% of October sequences). These findings demonstrate that our isolation with the culture-dependent technique captures the fine-scale genetic diversity of a subset of the diversity revealed by the culture-independent technique. With this picture in mind, we can better interpret the broader applicability of results stemming from each of these techniques.