Some isolates of the bacterium Pseudomonas fluorescens produce the antibiotic diacetylphloroglucinol (DAPG). DAPG is toxic to various organisms, including plants, fungi, viruses, and bacteria. In addition, crop yield increases have been reported after application of DAPG-producing isolates of P. fluorescens. The goal of this study was to determine whether DAPG is toxic to selected soil-dwelling nematodes, including free-living nematodes and plant parasites. In laboratory assays, the nematodes Caenorhabditis elegans, Heterodera glycines, Meloidogyne incognita, Pratylenchus scribneri, Pristionchus pacificus, Rhabditis rainai, and Xiphinema americanum were immersed in DAPG at concentrations ranging from 1 to 75 or 100 μg/ml. There were no observable effects on some species, but DAPG inhibited M. incognita egg hatch, tended to stimulate C. elegans egg hatch, and was toxic to X. americanum adults. The results indicate that DAPG produced by bacteria in the soil would not directly suppress population numbers of every nematode species exposed to the compound; effects would vary with nematode taxon and with life stage of the nematode.