An efficient technique for in vitro preservation of Agaricus subrufescens (=A. brasiliensis)

Agaricus subrufescens is one of the most important fungi produced biotechnologically in Brazil. After subculture, it shows a reduction in mycelial growth and difficulties with recovery. Hence, it is important to establish preservation and recovery techniques for its maintenance. Mycelium on a solid medium supplemented with activated charcoal (AC; technique A) was compared with culture under mineral oil (B) and mycelial disks in sterile water (C), all intended for storage of up to 12 months at 8°C. Mycelium viability, radial growth rate and genetic stability, assessed by random amplified polymorphic DNA (RAPD) analysis, were evaluated continuously. Technique A showed a 75% viability in the initial and final periods; technique B showed a 50% reduction in viability towards the end of the storage period; and technique C differed from the others insofar as viability was high until the 6th month, with a 50% drop after the 10th month, and a complete loss of viability after the 12th month. Overall, throughout the storage period, the rate of radial mycelium growth decreased. However, improved results were obtained when the media were also supplemented with AC. RAPD profiles demonstrated high genetic homogeneity of the mycelia maintained under techniques A, B and C (> 99%). In short, AC was shown to be an efficient ingredient for preservation within the 12-month storage period. Furthermore, it adds an important effect in that further recovery of A. subrufescens without significant morphological and genetic changes can be achieved.