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Mating behaviour and vegetative compatibility were studied in Spanish populations of Botryotinia fuckeliana. Fifty-seven isolates out of the 61 tested were sexually fertile with one or more of the reference strains of known mating type (MAT1-1 or MAT1-2). Thirty-nine isolates were heterothallic, giving fertile crosses when mated with the MAT1-1 (24 isolates) or the MAT1-2 (15 isolates) reference strain. Eighteen isolates crossed successfully with both MAT1-1 and MAT1-2 reference strains, and were referred to as homothallic or MAT1-1/2. Both mating types were widespread, being represented in isolates from two regions, from the same and different greenhouses, from different hosts, and from different years of isolation. Isolates were paired on Malt Extract Agar + NaCl to evaluate vegetative compatibility. Most of the paired isolates were unable to fuse and showed a different reaction of incompatibility. Nitrate-non-utilising (nit) mutants were selected by growth on a medium amended with 30–50gl−1 potassium chlorate. Over 600 chlorate-resistant sectors were recovered from 40 isolates at a mean frequency of 0.15–2.39 sectors per colony, but only 11% were identified as nit mutants by their thin growth with no aerial mycelium on minimal medium. However, most of these nit mutants reverted to wild type during six months of storage on chlorate-amended medium. Genetic complementation between nit mutants occurred only in two cases between mutants from the same isolate.