Extraction from Plant Tissue and Germination of Oospores of Peronospora viciae f.sp. pisi

A protocol was developed to extract oospores of Peronospora viciae f.sp. pisi from plant tissue and control bacterial contamination in a germination assay. Oospores were extracted by comminuting infected leaves and pods in water, sonicating the suspension and sieving it through mesh sizes 53 and 20 pm, respectively. Germination of oospores was negatively influenced by addition of chloramphenicol and penicillin. A combination of ampicillin and rifampicin strongly inhibited bacterial growth at 10 C, and did not negatively affect germination. Washing oospores in water or 0.02% Tween-80, and sonication did not influence germination. Treating oospore suspensions with cellulase buffered at pH 4.6 for 2 h digested most plant tissue but did not influence germination. Incubation in 0.05 m acetate buffer delayed germination. Germination was unaffected when oospores were incubated in 0.05 m citrate buffer.