Typing of anastomosis groups of Rhizoctonia solani by restriction analysis of ribosomal DNA

A method based on restriction analysis of polymerase chain reaction (PCR)-amplified ribosomal DNA was developed for the rapid characterization of large populations of Rhizoctonia solani at the anastomosis group (AG) level. The restriction maps of the internal transcribed spacers (ITS) sequences were compared for 219 isolates of R. solani belonging to AG-1 to AG-12 and AG-BI, representing diverse geographic and host range origins. Four discriminant restriction enzymes (MseI, AvaII, HincII, and MunI) resolved 40 restriction fragment length polymorphism (RFLP) types among the 219 ITS sequences of R. solani. Each RFLP type could be assigned to a single AG except for two RFLP types, which were common to two AG. A fifth enzyme allowed the discrimination of AG-6 and AG-12. In addition, the combination of four enzymes allowed the discrimination of subsets within AG-1, AG-2, AG-3, and AG-4. The efficiency of the typing method was confirmed by analyzing PCR-amplified ITS sequences of 30 reference strains. Furthermore, the PCR-RFLP method was used to characterize at the AG level 307 isolates of R. solani originating from ten sugar beet fields exhibiting patches of diseased plants in France. The PCR-based procedure described in this paper provides a rapid method for AG typing in R. solani.