Thirty-one soil samples from 14 different fields of Guatemala melon with vine decline symptoms were analyzed for the presence of organisms associated with the disease. With a soil-dilution plating method, only Macrophomina phaseolina was detected in five samples. With a melon bait plant technique, Olpidium bornovanus, often together with Melon necrotic spot virus (MNSV), was found in nearly all the samples, corresponding with all the fields studied. Other pathogens that were detected less frequently included Pythium aphanidermatum, Monosporascus cannonballus, and Rhizoctonia solani. Consequently, O. bornovanus and MNSV were uniquely associated with disease occurrence and thus are the most probable cause of melon vine decline in the fields studied.