Background: Section Calochroi is one of the most species-rich lineages in the genus Cortinarius (Agaricales, Basidiomycota) and is widely distributed across boreo-nemoral areas, with some extensions into meridional zones. Previous phylogenetic studies of Calochroi (incl. section Fulvi) have been geographically restricted; therefore, phylogenetic and biogeographic relationships within this lineage at a global scale have been largely unknown. In this study, we obtained DNA sequences from a nearly complete taxon sampling of known species from Europe, Central America and North America. We inferred intra-and interspecific phylogenetic relationships as well as major morphological evolutionary trends within section Calochroi based on 576 ITS sequences, 230 ITS + 5.8S + D1/D2 sequences, and a combined dataset of ITS + 5.8S + D1/D2 and RPB1 sequences of a representative subsampling of 58 species.

Phylogenetic relationships in the genus Cortinarius with emphasis on European species were studied by integrating a molecular phylogenetic analysis of nuclear rDNA sequences with morphological, chemical, and ecological data. Internal transcribed spacers (ITS1 and ITS2, including 5.8S) and the D1–D2 regions of nuclear rDNA of 262 Cortinarius species including most of the recognised subgenera from Europe, South America, Australia, Tasmania, New Zealand, and Asia were sequenced and analysed by maximum likelihood and neighbour joining. Our morphological and molecular analyses showed considerable disagreement with earlier morphology-based hypotheses concerning the infrageneric delimitations in Cortinarius. We propose a new base for a more natural classification system in Cortinarius involving the taxonomic rearrangement of the species into the following major lineages that we informally designate as “clades” at this time: (1) phlegmacioid species are included in Alluti, Amarescentes, Arguti, Calochroi, Caerulescentes, Percomes, Phlegmacioides, Phlegmacium, Purpurascentes, Scauri, and Vulpini; (2) the Heterocliti clade includes species of the subgenera Leprocybe and Cystogenes ; (3) most of the Myxacium species are included in the Delibuti and Myxacium clades; (4) part of Rozites are included in the Rozites clade; (5) the north-hemispheric species of subgenus Dermocybe are included in the Dermocybe clade and part of the south-hemispheric species in the Splendidi clade; (6) part of Sericeocybe are integrated in the Anomali and Telamonia clades; (7) members of subgenus Cortinarius are assigned to the Cortinarius clade; (8) telamonioid taxa are included in the Obtusi, Renidentes, and Telamonia clades. Several Cortinarius species with peculiar patterns of morphology and basidiome coloration had isolated positions in the sequence analysis, and the evolutionary relationships among the major lineages remained unresolved. Our results suggest the convergent evolution of certain morphological traits across phylogenetically divergent lineages in Cortinarius. However, combinations of features such as pileipellis anatomy, spore shape, veil coloration, basidiome pigments, and stipe shape appear useful to circumscribe clades. Our molecular phylogenetic analysis allows insights in global geographical patterns of species distribution within the major lineages in the genus Cortinarius. The clade structure proposed here could be used as backbone for further more detailed systematic studies.

Environmentally caused changes in chromosomes that do not alter the DNA sequence but cause phenotypic changes by altering gene transcription are summarized as epigenetics. A major epigenetic mechanism is methylation or demethylation at CpG-rich DNA islands. DNA methylation triggered by drugs has largely unexplored therapeutic consequences. Here we report increased methylation at a CpG rich island in the OPRM1 gene coding for μ-opioid receptors and at a global methylation site (LINE-1) in leukocytes of methadone-substituted former opiate addicts compared with matched healthy controls. Higher DNA methylation associated with chronic opioid exposure was reproduced in an independent cohort of opioid-treated as compared to non-opioid-treated pain patients. This suggests that opioids may stimulate DNA methylation. The OPRM1 methylation had no immediate effect on μ-opioid receptor transcription and was not associated with opioid dosing requirements. However, the global DNA methylation at LINE-1 was significantly correlated with increased chronic pain. This suggests inhibitory effects on the transcription of still unspecified nocifensive gene products. It further implies that opioids may be causally associated with increased genome-wide DNA methylation, although currently there is no direct evidence of this. This has phenotypic consequences for pain and may provide a new, epigenetics-associated mechanism of opioid-induced hyperalgesia. The results indicate a potential influence of opioid analgesics on the patients' epigenome. They emphasize the need for reliable and cost-effective screening tools and may imply that high-throughput screening for lead compounds in artificial expression systems may not provide the best tools for identifying new pain medications.