The study aimed to assess comparatively the accuracy and efficiency of three culture media protocols for estimating black-foot disease pathogens populations in soils and to examine how shifts in the abundance and composition of black-foot pathogens correspond to changes in specific soil properties. Glucose-Faba Bean Rose Bengal Agar (GFBRBA) was selected as the most efficient culture medium. All naturally infested soils tested positive for the presence of black-foot pathogens. Dactyonectria torresensis was the most frequently isolated species, followed by Dactylonectria alcacerensis and Ilyonectria liriodendri. A positive relationship between calcium carbonate and the Colony-Forming Units (CFUs) level of black-foot pathogens in soil was obtained. In this study, we provide an early, specific, and accurate detection of viable propagules of black-foot pathogens in soil, which is critical to understand the ecology of these fungi and to design effective management strategies.