Use of cryo-techniques with scanning electron microscopy to study infection of mature red raspberry fruits by Botrytis cinerea



Scanning electron microscopy (SEM) was used to study the infection of the mesocarp cells of ripe drupelets of the compound fruit of red raspberry (Rubus idaeus L.) by Botrytis cinerea Pers.: Fr. 24 h after the fruit had been wounded and inoculated with dry conidia. Frozen fully hydrated specimens showed that conidia had germinated in juice at the point of inoculation and in partially freeze-dried specimens hyphae were visible in some cells. However, structural detail was largely obscured by ice segregation zones in the closely packed large columnar, poyhedral cells of the fruit. These artifacts were removed in specimens which had been cryotrimmed and fully freeze-dried. In such specimens there were four distinct zones around the inoculation wound. Intercellular and intracellular hyphae were visible in the innermost area (zone 1); the walls of some infected cells were swollen, eroded and porous. In zone 2, an area of cells immediately beyond the hyphae, little structure was retained but in zone 3 cell walls were well preserved and showed lamellar or reticulate material in the intercellular spaces. The outer zone (zone 4) consisted of apparently healthy cells.