Despite their essential role in the environment, the number of known fungal species is still low compared to the recent estimates of the fungal biodiversity (from 2.2 to 3.8 million species), principally because of their often cryptic or ambiguous morphological traits. Recent studies have reported that only about approx. 35,000 correctly identified fungal species are represented by DNA sequences in public databases. This corresponds to a mere 1% of the estimated total number of species. Thus, the linkage of curated DNA sequence data to properly identified voucher specimens is of fundamental importance to fill the present gap between the different sizes of described and sequenced fungal diversity. For this purpose, mycological herbarium collections are considered an important source for fungal DNA-barcoding, and collection-based sequencing is a relevant priority for the coming decades. Unfortunately, ancient herbarium samples have both time and conservation related DNA damages, besides exogenous DNA contamination, that make nucleic acid extraction and amplification challenging. Here, we present the results of DNA extraction, ITS2 amplification and Illumina MiSeq sequencing of 36 specimens from the Saccardo Mycological Herbarium that were collected in the late XIX century and assigned to the genus Peziza. High-throughput sequencing was chosen as an alternative to the conventional Sanger- and cloning-based methods to overcome the high fragmentation of the ancient DNA and the massive occurrence of non-target DNA from fungal contaminants. Our approach has permitted the assignment of ITS2 sequences to 23 out of the 36 specimens studied in this work, thus providing a univocal DNA sequence for those one century old samples. Furthermore, the ITS2 sequence analysis has permitted a taxonomic study of the samples that has resulted in a re-evaluation of five samples at the species level and 18 samples at genus or higher level. Our results highlight the possibility to apply the technique presented in this work also to the old and more precious type specimens in order to relate a DNA sequence to these important samples, coupling the traditional morphological description of the species with a DNA sequence.

In a recent monograph on the genus Rosellinia, type specimens worldwide were revised and re-classified using a morphological approach. Among them, some came from Pier Andrea Saccardo’s fungarium stored in the Herbarium of the Padova Botanical Garden. In this work, we taxonomically re-examine via a morphological and molecular approach nine different Roselliniasensu Saccardo types. ITS1 and/or ITS2 sequences were successfully obtained applying Illumina MiSeq technology and phylogenetic analyses were carried out in order to elucidate their current taxonomic position. Only the ITS1 sequence was recovered for Rosellinia areolata, while for R. geophila, only the ITS2 sequence was recovered. We proposed here new combinations for Rosellinia chordicola, R. geophila and R. horridula, while for R. ambigua, R. areolata, R. australis, R. romana and R. somala, we did not suggest taxonomic changes compared to the current ones. The name Rosellinia subsimilis Sacc. is invalid, as it is a later homonym of R. subsimilis P. Karst. & Starbäck. Therefore, we introduced Coniochaeta dakotensis as a nomen novum for R. subsimilis Sacc. This is the first time that these types have been subjected to a molecular study. Our results demonstrate that old types are an important source of DNA sequence data for taxonomic re-examinations.

A new species, Leucoagaricus cupresseoides, from a pine forest in Castelfusano, Rome, Italy, is described based on both morphological and molecular analyses. Updated morphological descriptions of Leucoagaricus aurantiovergens and L. pseudopilatianus are presented due to new collections from Italy and confirmed by nrITS sequence analysis of the type specimens of the two species. Based on our morphological and molecular analyses, L. pseudopilatianus var. roseodiffractus and L. paracupresseus are synonyms of L. aurantiovergens; and L. pseudopilatianus var. rugosoreticulatus is not molecularly distinct from L. pseudopilatianus. Leucoagaricus cupresseus, L. cupresseoides, L. aurantiovergens and L. pseudopilatianus form the L. cupresseus complex for which a comparative table is provided.

A new species of Xylaria is described based on morphological characters of both sexual and asexual morphs, and molecular data based on nuclear rDNA internal transcribed spacer, α-actin, β-tubulin and RNA polymerase subunit II sequences. Xylaria aurantiorubroguttata is characterized by the presence of both upright, cylindrical, long-stipitate and globose to subglobose, short-stipitate stromata, immature stromatal stages producing at first orange and then red drops, and ascospores with a slightly oblique, straight half spore-length germ slit. We provide also new morphological descriptions for X. haemorrhoidalis (holotype) and X. anisopleura (isosyntype), two Xylaria species belonging to X. polymorpha complex together with X. aurantiorubroguttata.

Specimens of Candolleomyces from tropical and subtropical regions, mainly the Dominican Republic, the Caribbean, Central America, and southern USA, were analyzed morphologically and molecularly. Historical fungarium types and sequences from Africa, Asia, and the Americas were included. ITS sequences from ancient collections were obtained via Illumina when Sanger sequencing failed. Phylogenetic analyses (Bayesian and Maximum Likelihood) of ITS and, where available, tef1-α clarified poorly known species and revealed new synonymies. Six species are described as new: C. cingulatus, C. dennisianus, C. djondjon, C. fuscodiscus, C. peglerianus, and C. subannulatus; two taxa (C. luteidiscus and C. singerianus) are proposed as nomina provvisoria. Twenty-seven new combinations are proposed, and several historical taxa are synonymized. Only four species occur across both the American and the African/Asian tropics, indicating that climate-selective distribution and regional isolation are drivers of speciation. Ethnomycological notes on the Haitian culinary dish of djon-djon are provided, and a dichotomous key to tropical American and African species is included, refining taxonomy and revealing previously overlooked diversity.