2.6 Antagonistic efficacy of the selected strain of
R. mucilaginosa
The antagonistic efficacy of the selected strain of R. mucilaginosa (RKZ1-5) to P. expansum was evaluated on pears. The procedure was similar to that described in Section 2.4. Six wounds were made on each pear with a sterile cork-borer. R. mucilaginosa suspension (50 μl) in sterile distilled water at 1×105, 1×106, 1×107, 1×108, or 1×109 cells/ml were pipetted into each wound, respectively. The same volume of sterile distilled water was pipetted into wounds to be used as controls. After 3 h, 30 μl of pathogen spores at 1×104 cells/ml were inoculated into the wounds.
The pear samples were incubated as described in Section 2.4. The biocontrol activity was assessed 6 d after treatment. Each yeast treatment consisted of three replicates, each consisting of 12 fruits. The experiment was conducted twice.
2.7 Antagonistic action of different preparations of R. mucilaginosa to P. expansum on pears
The pear fruits were wounded using the proce- dure described in Section 2.4. Based on the results of the experiments (Section 2.6), the most effective cell concentration of R. mucilaginosa antagonistic to P. expansum was 1×108 cells/ml and this was used in further tests based on the following preparations: (1) 1×108 cells/ml unwashed cell culture mixture (i.e. the yeast cells in the nutrient yeast broth medium); (2) 1×108 cells/ml washed cell suspension (prepared as described in Section 2.4); (3) cell-free culture fil- trate (made by filtering the supernatant of the centri- fuged yeast culture medium through 0.22 μm sterile Millipore filtration); (4) autoclaved cell culture (the R. mucilaginosa culture was sterilized by moist heat sterilization at 121 °C for 20 min); (5) Sterile distilled water was used as a control. After air-drying, the treated pear samples were stored as described in Section 2.4. Decay incidence and lesion diameter were recorded after 6 d. Each yeast treatment con- sisted of three replicates, each consisting of 12 fruits. The whole experiment was conducted twice.