Genotypic diversity among Brazilian isolates of Sclerotium rolfsii

Thirty isolates of Sclerotium rolfsii Sacc. from different hosts and region s of Brazil were studied in relation to morphology, mycelial compatibility, analysis of genomic DNA through random amplified polymorphic DNA (RAPD), v ariation within the nuclear rDNA [internal transcribed spacers (ITS)] and s equencing of ITS fragments. There was considerable variability among isolat es in relation to the number, size and location of sclerotia on the medium surface. Thirteen mycelial compatibility groups (MCG) were identified among 23 isolates. Seven isolates were only self-compatible. With the exception of group 3, where all the isolates came from soybean, there was no apparent correlation between group and isolate origin. On the basis of RAPD profile s, 11 haplotypes (A to K) were identified. There was an association between the RAPD groups and MCG. Haplotypes A, B, D, G, I and K belonged to MCG gr oups 1, 2, 3, 4, 5 and 6, respectively. All other RAPD haplotypes contained incompatible isolates. Polymerase chain reaction (PCR) amplification with primers 4R and 5F amplified two fragments containing ITS1, ITS2 and 5.8 S r DNA sequences, that were present in all isolates, with molecular sizes of 7 39 and 715 bp. Restriction analysis of PCR products showed that the two fra gments had sequence divergency which is referred to as 'ITS types'. Four ar bitrarily chosen soybean isolates (2, 6, 7 and 23) and two non-soybean isol ates (11 and 22) were used to investigate the variation within the ITS sequ ence and its role in the phylogeny. The strict consensus of nine most-parsi monious trees inferred from the data set which included six isolates of S. rolfsii, four of which have two different 'ITS types', showed three well-su pported groupings. The neighbour-joining tree inferred from the data set al so showed three major clades as did the parsimony tree. The major differenc e was that in the neighbour-joining tree the 'ITS type' 11 was resolved and grouped in one clade. These results show that the 'ITS types' within isola tes are almost always phylogenetically distinct. There was no clear correla tion between ITS-based phylogeny and isolate origin.