Direct examination of soil for sporangia of Synchytrium endobioticum using chloroform, calcium chloride and zinc sulphate as extraction reagents

Fields infested with Synchytrium endobioticum can be descheduled when the soil is found free from sporangia of S. endobioticum . For direct examination, EPPO Standard PM 3/59 describes a soil extraction technique based on the use of a sieve shaker with six sieves. We compared recovery of sporangia between this (modified) method and an extraction method employed by the Dutch Plant Protection Service (PPS method). Recovery was determined using an inoculum dilution series: 125, 25, 5, 1, 0.2 or 0.04 sporangia per g soil. Extraction reagents used were chloroform and calcium chloride in the method described by EPPO, calcium chloride and zinc sulphate in the PPS method. At 125 sporangia per g soil, the mean density determined for the modified EPPO method was 228 sporangia per g soil when chloroform was used. Using calcium chloride, recovery percentage was higher for the modified EPPO method than for the PPS method (286, 136%, n.s. P  < 0.05). The advantage of the modified EPPO method was the larger soil volume to be processed; its disadvantages were use of complex equipment and noxious reagents (chloroform). Both extraction methods showed high variation in recovery between samples, making accurate estimation of sporangial densities in soil awkward.