Use of single-protoplast isolates in the study of the mating phenomena of Rhizoctonia solani (Thanatephorus cucumeris) AG-1 IC and IA

This study evaluates the effectiveness of using single-protoplast isolates (SPIs) to study the mating phenomena of Rhizoctonia solani AG-1 IC and IA. SPIs obtained from three field isolates (F-1, Rh28, and RO2) of AG-1 IC were paired with representative single-basidiospore isolate (SBI)-M1/-M2 testers, each from their own field isolates, or paired in all possible combinations. Tufts were formed between SPIs and SBI-M1/-M2 testers and between SPIs-M1 and -M2. The separation ratios of SPIs-M1 and -M2 were approximately 1 : 1, which were similar to the results obtained with SBIs. SPIs obtained from three isolates (GNSD, R59, and Tr8) of AG-1 IA, which failed to form basidiospores, were paired in all possible combinations. Although no tufts formed among SPIs from Tr8 and R59, tufts did form between SPIs from GNSD. SPIs from GNSD were separated into homokaryotic (-M1 or -M2) and heterokaryotic isolates, and the separation ratio of -M1 and -M2 was also around 1 : 1. Amplified fragment length polymorphism (AFLP) phenotypes of the tuft isolates formed between GNSD SPIs-M1 and -M2 suggested that these tuft isolates were all heterokaryotic. These results indicate that all three isolates of AG-1 IC and one isolate GNSD of AG-1 IA are heterokaryotic, and that the other two isolates of Tr8 and R59 of AG-1 IA are homokaryotic. Single-protoplast isolates are effective for studies of the mating phenomena of isolates belonging to different AGs of R. solani that could not form a perfect stage.