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Variation within the internal transcribed spacer (ITS) of the ribosomal RNA gene of 15 isolates representing seven European Armillaria species, was examined by sequencing of the PCR-amplified products. The analysis of an 744-bp region showed that the 5.8S gene appeared to be highly conserved in the 15 isolates and in other Basidiomycetes and Ascomycetes, whereas ITS1 and especially ITS2 spacers exhibited polymorphisms due to base substitutions, insertions or deletions of up to eight nucleotides. An initial dendrogram for the full sequence was drawn using cluster analysis (UPGMA), and a tree was constructed using the maximum parsimony method. Both methods indicated that the isolates could be divided into four major groups. One group, consisting of A. ectypa, was distinct from all the other species. Examination of the other groups indicated that A. tabescens and A. mellea were in a separated cluster, with a significant variation between the two isolates of the latter species. A. gallica and A. cepistipes constituted another closely related group distinguishable from A. ostoyae and A. borealis, these latter two species exhibiting the highest similarity. The results are consistent with, and discussed in regard to, the relationships estimated previously by pairing tests, morphological and physiological comparisons, as well as by restriction fragment length polymorphism of the rDNA.Key words : Armillaria, biological species, direct sequencing, internal transcribed spacer, phylogeny, ribosomal DNA. The genus Armillaria (Fr. : Fr.) Staude includes serious pathogens causing root rot of a broad host range of plants, all over the world. In Europe, there are seven intersterile groups or biological species (Korhonen, 1978 ;

Thirteen Armillaria isolates, collected from various geographical areas in tropical Africa and previously characterized by cultural morphology, pairing tests and isozyme analysis, were evaluated using the polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP). DNA regions corresponding to the intergenic spacer (IGS) and internal transcribed spacer (ITS) were amplified and analyzed by restriction enzyme digestion. The IGS amplification products were about 875 by long and uniform in length among the isolates. The amplified-ITS region showed two different lengths corresponding to two groups. The first group included the isolates believed to belong to A. mellea ssp. africana and two Kenyan isolates (K11 and K12) belonging to a yet unnamed biological species. The second group included isolates identified as A. heimii and a Tanzanian isolate (T7). Each length variant of the ITS showed distinct RFLP banding patterns. Digestion with EcoRI confirmed the two polymorlphic groups while the endonucleases Alul and Ndell discriminated the A. mellea isolates from the Kenyan isolates K11 and K12. In addition, the latter enzyme showed a slight dissimilarity between the A. heimii isolates from Western and Eastern Africa (C1 and Z1). Digestion with Hinfl cleaved the isolates of A. heimii into two sub-groups corresponding to the heterothallic and homothallic forms. This endonuclease also indicated that the isolate T7, originating from Tanzania, was clearly similar to the heterothallic species A. heimii. Data presented support the maintenance of three distinct species of Armillaria in tropical Africa with A. heimii as a variable species, the isolates of which were separated in accordance with their sexual system. The results indicate that PCR-RFLP can be used as a simple and speedy taxonomical tool for the ecological studies of Armillaria species. Armillaria is a genus of root-infecting fungi with worldwide distribution consisting of a number of distinct species. The number of these species in the world is about 30. Taxonomical studies of African Armillaria have included a wide range of investigation techniques such as observations of cultural characteristics (vegetative mat morphology, production of subterranean rhizomorphs, morphology of fruit bodies, and monospore isolates obtained in vitro) and pairing tests