Many small (temporary) collections of fungi maintained by plant pathologists during their research receive inadequate attention to ensure stability. Maintaining collections of fungi in pure and viable conditions, minimising physiological and morphological changes is, however, a necessity. The objective of this study was to find preservation techniques for three Rosellinia isolates used in our plant pathogenic research. Various inert and nutritious carriers, solid as well as liquid, were used to test their suitability for conserving these Rosellinia isolates. Different cryoprotectants, cooling rates and thawing rates were tested to optimise liquid nitrogen storage procedures. Survival and/or growth rate were assessed over time. Rosellinia bunodes was the most difficult to store with survival not exceeding six to nine months using traditional storage methods in mineral oil and silica gel. Storage of Rosellinia necatrix and Rosellinia pepo was successful for periods up to at least 16 months in several carriers and for up to two years for R. necatrix in silica gel. Storage in liquid nitrogen proved no problem for R. necatrix or R. pepo with a 100% survival in all cases, although radial growth rates after recuperation were affected by cryoprotectant and thawing rates. Storage of R. bunodes was more difficult and survival as well as growth rates were affected by cryoprotectant and thawing rates. Cooling rates did not affect radial growth in any of the isolates. The results showed that development of a generalised procedure for storage of our Rosellinia species was not possible and successful storage protocols had to be developed for individual isolates.