Comparison between Tuber gennadii and T. oligospermum lineages reveals the existence of the new species T. cistophilum (Tuberaceae, Pezizales).

New collections of T. gennadii, T. oligospermum and T. asa (Tuberaceae, Pezizales) from the Mediterranean basin were compared on the basis of morphology and DNA sequences. Phylogenetic inference based on the internal transcribed spacer region (ITS) and the 28S nuclear large ribosomal subunit (nrLSU) indicates that T. gennadii includes at least two lineages with characteristic morphological traits. T. gennadii s. str. has small subglobose ascomata with conspicuous glebal locules that form a continuous labyrinth when mature and has a prosenchymatic peridium. T. lacunosum has medium to large, deeply lobed ascomata with small to very small isolated locules when mature and has a pseudoparenchymatic peridium. Two major clades were identified within the T. oligospermum group. On the basis of morphological, ecological and biogeographical data, these are identified with the taxa T. oligospermum (= T. asa) and T. sphaerospermum. Tuber oligospermum forms small to medium globose or subglobose, whitish ascomata, with globose to broadly ellipsoidal ascospores and is found in alkaline, sandy soils. Tuber sphaerospermum forms medium to large, irregularly globose or deeply lobed, yellowish ascomata, with regularly globose ascospores and is found in acidic soils. Finally, a new truffle species with a prosenchymatic peridium and globose to ellipsoid reticulate spores, collected in acidic argillaceous soils of Spain under Cistus ladanifer is proposed here as the new species T. cistophilum.