We determined whether isolates of the take-all pathogen Gaeumannomyces graminis var. tritici become less sensitive to 2,4-diacetylphloroglucinol (2,4-DAPG) during wheat monoculture as a result of exposure to the antibiotic over multiple growing seasons. Isolates of G. graminis var. tritici were baited from roots of native grasses collected from noncropped fields and from roots of wheat from fields with different cropping histories near Lind, Ritzville, Pullman, and Almota, WA. Isolates were characterized by using morphological traits, G. graminis variety-specific polymerase chain reaction and pathogenicity tests. The sensitivity of G. graminis var. tritici isolates to 2,4-DAPG was determined by measuring radial growth of each isolate. The 90% effective dose value was 3.1 to 4.4 microg ml(-1) for 2,4-DAPG-sensitive isolates, 4.5 to 6.1 microg ml(-1) for moderately sensitive isolates, and 6.2 to 11.1 microg ml(-1) for less sensitive isolates. Sensitivity of G. graminis var. tritici isolates to 2,4-DAPG was normally distributed in all fields and was not correlated with geographic origin or cropping history of the field. There was no correlation between virulence on wheat and geographical origin, or virulence and sensitivity to 2,4-DAPG. These results indicate that G. graminis var. tritici does not become less sensitive to 2,4-DAPG during extended wheat monoculture.