Experimental Outcrossing in Agaricus bisporus Revealed a Major and Unexpected Involvement of Airborne Mycelium Fragments

Agaricus bisporus var. bisporus, the button mushroom, has a predominantly pseudohomothallic life cycle. Most of its spores are heterokaryotic and give rise to fertile heterokaryons. However, previous studies have suggested that outcrossing should not be rare in wild populations. In order to discover how outcrossing occurs, we experimentally favored it between aerial propagules of a fruiting donor mycelium and a delayed receiver mycelium that only invaded culture trays. Two donor/receiver pairs were studied, and potentially hybrid basidiomata collected on the receiver trays were analyzed with a mitochondrial marker, two unlinked nuclear CAPS markers, then haplotype markers based on DNA sequences obtained after PCR cloning of the rDNA ITS region and the fruk gene. For one of the two pairs, most basidiomata were hybrids between the donor and the receiver. Genotyping of the hybrids revealed only two genotypes consistent with outcrossing involving airborne mycelium fragments rather than basidiospores. The resident receiver heterokaryon that provided its mitochondria to the hybrid basidiomata is suspected to have had a trophic contribution to their growth and successful fruiting. The high level of heterozygosity and the cultivar introgression previously revealed in wild populations of this pseudohomothallic species may result from outcrossing involving airborne pieces of mycelium.