Heterobasidion annosum 5.8s ribosomal DNA and internal transcribed spacer sequence: rapid identification of European intersterility groups by ribosomal DNA restriction polymorphism

Abstract

Using conserved fungal ribosomal gene sequences the internal transcribed spacer (ITS) regions one and two (ITS1, ITS2) and the 5.8s ribosomal RNA gene (rRNA) of Heterobasidion annosum were amplified by the polymerase chain reaction (PCR). The nucleotide sequence was determined in three European intersterility groups (ISG-S, -F and -P). Three sequence variants of the ITS were found in ISG-S isolates. The sequence of the ITS of ISG-F differed by two residues from the major ISG-S sequence variant. The ISG-P sequence differed from ISG-S and ISG-F at 15-16 and 16 residues, respectively. Amplified intergenic spacer elements were informative for ISG fingerprinting following digestion with various 4-cutter restriction endonucleases. All differences in the restriction fragments between the ISGs were because of sequence differences in the ITS regions. The fingerprint patterns of isolates from the same intersterility group but from different European localities were identical. These results show that ribosomal DNA fingerprinting is a rapid technique to identify ISGs in Heterobasidion annosum.