Basidiomes of a Pisolithus species were collected from a ca 2500 m2 Australian sclerophyll forest site during 1997 and 1999 and of a second Pisolithus species from a further ca 150 m2 site during 1999. Inter-simple sequence repeat (ISSR) PCR was conducted on DNA extracted from each basidiome using the primers 5′BDB(ACA)5, 5′DDB(CCA)5 and 5′DHB(CGA)5. Thirty-seven genotypes of Pisolithus species I were detected at the North Wilberforce site, with eight genotypes present during both 1997 and 1999. All other genotypes were observed during only one year. Mapping genotype distribution according to location of basidiome collection at the site suggested that most genotypes were present as small (< 2·0 m2) below-ground mycelial genets, however several larger (> 4·0 m2) genets (including one ca 120 m2 genet) were also present. All Pisolithus species II basidiomes at the North Turramurra site were of a common genotype, suggesting the presence of a single large (ca 69 m2) below-ground mycelial genet of this taxon at the site. Both Pisolithus species thus appear to produce large long-lived soil-borne mycelia, but establishment of large genets may be restricted under some circumstances.