Water Sample Filtration and Nucleic Acid Extraction

Prior to the experiment, sample collection bottles were soaked in 10% sodium hypochlorite and repeatedly rinsed in water. After the experiment, approximately 300 mL from each sample bottle was filtered through a 0.8-μm cellulose nitrate filters of 25-mm diameter (Whatman item number 7188-002). Filtration occurred under 30 psi negative pressure (Thomas Industries model 907CA18-554 air compressor) using standard filtration flasks and fittings. The filtering apparatus was soaked in 10% sodium hypochlorite between uses, washed twice with tap water and once with deionized water, then air-dried. Filters were handled with ethanol-sterilized forceps. Processed filters were folded into a 15-mL Falcon tube (Corning) and stored in a freezer rated at −20 °C (operational temperature was measured to be approximately −12 °C). Filters were later transported on ice to the US Geological Survey, Leetown Science Center, Kearneysville, West Virginia, for nucleic acid extraction with the cetyl trimethylammonium bromide protocol of Renshaw et al. (2015) (link). Extraction yield was measured with a Qubit fluorometer using the High-sensitivity DNA kit (ThermoFisher, Waltham, Massachusetts, USA).