Enrichment cultures and isolation of guanylurea-degrading bacteria.

The bacterial Pseudomonas mendocina strain GU was isolated from a sample of activated sludge collected at the metropolitan wastewater treatment plant in Saint Paul, MN. Isolation was achieved by enrichment culture with citrate-acetate medium and 1 g of sludge per 50 ml of minimal medium as the inoculum. The minimal medium contained the following per liter of deionized water: 5.45 g K₂HPO₄, 0.2 g MgSO₄·7H₂O, 0.1 g NaCl, 1.5 g 20 mM sodium acetate, and 20 mM sodium citrate (54). Guanylurea (1 mM) was then added as the sole nitrogen source. Enrichments and isolates were grown at 30°C in a shaking incubator at 200 to 225 rpm. Cultures were transferred in 10-fold dilutions into fresh medium every 7 days. Individual isolates were obtained by plating 10-fold serial dilutions of the enrichments on selective guanylurea plates, transferred to Luria-Bertani (LB) plates, and then isolated by streaking on LB and minimal medium plus guanylurea plates until pure.