Microbiome of observed in experimental laboratory microcosms exposed to variable nitrapyrin and nitrogen amendments in the presence or absence of nitrifier enhanced microbial community
The NEM1 culture was derived from a sediment microbiome collected from a golf course pond in the Columbia, Missouri. Triplicate sediment samples were collected with sterile spatulas into sterile 50 ml capped plastic tubes. To select for ammonia oxidizing microorganisms, 1 g of sediment was transferred into 50 ml of autoclaved mineral salts minimal media, enriched with 2 mg N/L total ammonia nitrogen (TAN, added as ammonium sulfate), and hereafter referred to as the media (media recipe from Verhagen and Laanbroek, 1991). Sediment cultures were prepared in triplicate 250 ml Erlenmeyer flasks capped with sterile foam stoppers or aluminum foil and incubated in a dark incubator at 22C. Cultures were monitored for ammonia depletion and nitrite and nitrate production using HACH TNT plus 830 ammonia kits, 839 nitrite kits, and 835 nitrate kits, respectively (HACH Company, Loveland, CO). Samples that showed consistent ammonia consumption trends (i.e., 80% of ammonia depleted in no more than 7 days) were transferred to new media weekly, using a ratio of 1:10 culture to new media volume. After approximately four weeks, a stable, consistent ammonia consumption rate of about 1.6 mg/L ammonia-nitrogen per week was achieved and culture volumes were increased from 50 ml to 100 ml.