Ester Gaya

Implementing a cumulative supermatrix approach for a comprehensive phylogenetic study of the Teloschistales (Pezizomycotina, Ascomycota)

The resolution of the phylogenetic relationships within the order Teloschistales (Ascomycota, lichen-forming-fungi), with nearly 2000 known species and outstanding phenotypic diversity, has been hindered by the limitation in the resolving power that single-locus or two-locus phylogenetic studies have provided to date. In this context, an extensive taxon sampling within the Teloschistales with more loci (especially nuclear protein-coding genes) was needed to confront the current taxonomic delimitations and to understand evolutionary trends within this order. Comprehensive maximum likelihood and bayesian analyses were performed based on seven loci using a cumulative supermatrix approach, including protein-coding genes RPB1 and RPB2 in addition to nuclear and mitochondrial ribosomal RNA-coding genes. We included 167 taxa representing 12 of the 15 genera recognized within the currently accepted Teloschistineae, 22 of the 43 genera within the Physciineae, 49 genera of the closely related orders Lecanorales, Lecideales, and Peltigerales, and the dubiously placed family Brigantiaeaceae and genus Sipmaniella. Although the progressive addition of taxa (cumulative supermatrix approach) with increasing amounts of missing data did not dramatically affect the loss of support and resolution, the monophyly of the Teloschistales in the current sense was inconsistent, depending on the loci-taxa combination analyzed. Therefore, we propose a new, but provisional, classification for the re-circumscribed orders Caliciales and Teloschistales (previously referred to as Physciineae and Teloschistineae, respectively). We report here that the family Brigantiaeaceae, previously regarded as incertae sedis within the subclass Lecanoromycetidae, and Sipmaniella, are members of the Teloschistales in a strict sense. Within this order, one lineage led to the diversification of the mostly epiphytic crustose Brigantiaeaceae and Letrouitiaceae, with a circumpacific center of diversity and found mostly in the tropics. The other main lineage led to another epiphytic crustose family, mostly tropical, and with an Australasian center of diversity--the Megalosporaceae--which is sister to the mainly rock-inhabiting, cosmopolitan, and species rich Teloschistaceae, with a diversity of growth habits ranging from crustose to fruticose. Our results confirm the use of a cumulative supermatrix approach as a viable method to generate comprehensive phylogenies summarizing relationships of taxa with multi-locus to single locus data.

Phylogenetic study of Fulgensia and allied Caloplaca and Xanthoria species (Teloschistaceae, lichen-forming ascomycota)

Fulgensia Massal. & De Not. is a widespread genus with considerable morphological and ecological heterogeneity across species. For this reason, the taxonomic delimitation of this genus has been controversial. Relationships among species of Fulgensia, Caloplaca Th. Fr., and Xanthoria (Fr.) Th. Fr. (Lecanorales) were investigated based on a comprehensive phylogenetic analysis of 62 DNA sequences from the nuclear ribosomal internal transcribed spacer (ITS) region using maximum parsimony (MP) and likelihood (ML). Ambiguously aligned (INAASE coded characters) and unambiguous regions were analyzed separately and combined when using MP as the optimization criterion. All our analyses confirm the polyphyly of this genus as three distinct lineages: Fulgensia sensu stricto, F. australis, and F. schistidii. We report here that Caloplaca, Fulgensia, and Xanthoria together form two main sister lineages. One lineage includes Fulgensia schistidii (part of the C. saxicola group), Xanthoria, and most of the lobed Caloplaca species belonging to the Gasparrinia group. A second main lineage comprises the remaining Caloplaca species, Fulgensia sensu stricto, and F. australis. Therefore, the traditional generic level classification schemes for the family Teloschistaceae appear to be highly artificial. All three genera were found to be nonmonophyletic. We demonstrate here that the ITS is appropriate to resolve relationships across the Teloschistaceae. However, a combination of an MP analysis, in which ambiguously aligned regions are accommodated using INAASE, with an ML analysis, in which phylogenetic confidence is estimated using a Bayesian approach, is needed.

Phylogenetic reassessment of the Teloschistaceae (lichen-forming Ascomycota, Lecanoromycetes).

The Teloschistaceae is a widespread family with considerable morphological and ecological heterogeneity across genera and species groups. In order to provide a comprehensive molecular phylogeny for this family, phylogenetic analyses were carried out on sequences from the nuclear ribosomal ITS region obtained from 114 individuals that represent virtually all main lineages of Teloschistaceae. Our study confirmed the polyphyly of Caloplaca, Fulgensia and Xanthoria, and revealed that Teloschistes is probably non-monophyletic. We also confirm here that species traditionally included in Caloplaca subgenus Gasparrinia do not form a monophyletic entity. Caloplaca aurantia, C. carphinea and C. saxicola s. str. groups were recovered as monophyletic. The subgenera Caloplaca and Pyrenodesmia were also polyphyletic. In the subgenus Caloplaca, the traditionally recognized C. cerina group was recovered as monophyletic. Because this study is based solely on ITS, to maximize taxon sampling, the inclusion of phylogenetic signal from ambiguously aligned regions in MP (recoded INAASE and arc characters) resulted in the most highly supported phylogenetic reconstruction, compared with Bayesian inference restricted to alignable sites.

Align or not to align? Resolving species complexes within the Caloplaca saxicola group as a case study

The Caloplaca saxicola group is the main group of saxicolous, lobed-effigurate species within genus Caloplaca (Teloschistaceae, lichen-forming Ascomycota). A recent monographic revision by the first author detected a wide range of morphological variation. To confront the phenotypically based circumscription of these taxa and to resolve their relationships morphological and ITS rDNA data were obtained for 56 individuals representing eight Caloplaca species belonging to the C. saxicola group. We tested the monophyly of these eight morphospecies by performing maximum parsimony, maximum likelihood and two different types of Bayesian analyses (with and without a priori alignments). Restricting phylogenetic analyses to unambiguously aligned portions of ITS was sufficient to resolve, with high bootstrap support, five of the eight previously recognized species within the C. saxicola group. However, phylogenetic resolution of all or most of the eight species currently included as two distinct subgroups within the C. saxicola group was possible only by combining morphological characters and signal from ambiguously aligned regions with the unambiguously aligned ITS sites or when the entire ITS1 and 2 regions were not aligned a priori and included as an integral component of a Bayesian analysis (BAli-Phy). The C. arnoldii subgroup includes C. arnoldii, comprising four subspecies, and the C. saxicola subgroup encompasses seven species. Contrary to the C. saxicola subgroup, monophyly of taxa included within the C. arnoldii subgroup and their relationships could not be resolved with combined ITS and morphological data. Unequivocal morphological synapomorphies for all species except C. arnoldii and C. pusilla are recognized and presented.

Expansion of the Stictidaceae by the addition of the saxicolous lichen-forming genus Ingvariella

The monotypic, lichen-forming genus Ingvariella originally was segregated from Diploschistes and placed within the Thelotremataceae (Ostropales) based on aspects of exciple morphology. However, the I+ hymenium and amyloid ascus wall suggest affinities to families other than the Thelotremataceae. To assess the identity of Ingvariella and to investigate its placement within the Ostropales, we inferred phylogenetic relationships of I. bispora by comparison of mtSSU rDNA and nuLSU rDNA sequences for 59 species encompassing a broad array of ostropalean fungi by means of Bayesian, maximum likelihood and weighted maximum parsimony methods. Here we report that Ingvariella is a member of the Stictidaceae, sister to the mainly saprotrophic genus Cryptodiscus. The inclusion of the first saxicolous lichen-forming fungus within this family expands the broad ecological diversity of the Stictidaceae, where saprotrophic fungi, corticicolous lichen-forming fungi and lichenized and non-lichenized conspecific taxa have been described previously. We also present new insights into the relationships among other families within the Ostropales.

The adaptive radiation of lichen-forming Teloschistaceae is associated with sunscreening pigments and a bark-to-rock substrate shift

Significance The tempo of diversification of life can be accelerated by fortuitous ecological opportunity or by phenotypic innovation. In this study, we document how both factors are likely to have played a role in the origin and success of a major fungal lineage, the Teloschistaceae (comprising ∼1% of all fungi). Anthraquinone pigments are found in a widespread, but scattered, range of fungi and plants, but are particularly abundant in the lichen-forming Teloschistaceae, where they provide sunlight protection, especially needed when growing in arid deserts of the world. We found that anthraquinones evolved in these lichens, in conjunction with an ecological switch to exposed, rocky environments, allowing them to colonize swathes of unexploited habitats worldwide and sparking an acceleration in diversification. Adaptive radiations play key roles in the generation of biodiversity and biological novelty, and therefore understanding the factors that drive them remains one of the most important challenges of evolutionary biology. Although both intrinsic innovations and extrinsic ecological opportunities contribute to diversification bursts, few studies have looked at the synergistic effect of such factors. Here we investigate the Teloschistales (Ascomycota), a group of >1,000 lichenized species with variation in species richness and phenotypic traits that hinted at a potential adaptive radiation. We found evidence for a dramatic increase in diversification rate for one of four families within this order—Teloschistaceae—which occurred ∼100 Mya (Late Cretaceous) and was associated with a switch from bark to rock and from shady to sun-exposed habitats. This adaptation to sunny habitats is likely to have been enabled by a contemporaneous key novel phenotypic innovation: the production in both vegetative structure (thallus) and fruiting body (apothecia) of anthraquinones, secondary metabolites known to protect against UV light. We found that the two ecological factors (sun exposure and rock substrate) and the phenotypic innovation (anthraquinones in the thallus) were all significant when testing for state-dependent shifts in diversification rates, and together they seem likely to be responsible for the success of the Teloschistaceae, one of the largest lichen-forming fungal lineages. Our results support the idea that adaptive radiations are driven not by a single factor or key innovation, but require a serendipitous combination of both intrinsic biotic and extrinsic abiotic and ecological factors.

Twenty-five cultures of lichenizing fungi available for experimental studies on symbiotic systems

In this study we describe the techniques used to culture 25 mycobionts spanning three classes and five orders of the leotiomyceta (Ascomycota). We find that five media, including potato-carrot, malt extract-yeast extract (MY), Bold’s basal medium with nitrogen (NMBBM), oatmeal, and yeast extract with supplements (YES), are sufficient to induce ascospore germination of many lichenizing fungi and are also suitable for maintaining growth of the culture over the long term. Regular physical disruption of the cultures in liquid media is recommended to stimulate continued growth. Genomes of five of these lichen-forming fungal strains have been sequenced. The identity of each culture was confirmed by sequencing the nuclear ribosomal internal transcribed spacer (ITS) or the mitochondrial small subunit (mitSSU) from each strain. Additionally, the level of sequencing in terms of total number of genes sequenced for each taxon is provided. All fungal cultures have been deposited in public culture collections and, therefore, are available to the scientific community for conducting in vitro experiments.

Phylogenetic study of Diploschistes (lichen-forming Ascomycota: Ostropales: Graphidaceae), based on morphological, chemical, and molecular data

The genus Diploschistes includes crustose lichen-forming fungi with a carbonized proper excipulum with lateral paraphyses, and a chemistry dominated by orcinol depsides. However, the taxon D. ocellatus lacks these excipular characters and has β-orcinol depsidones, raising doubts about its inclusion within this genus. Using a two-locus dataset (mtSSU, nuLSU), our phylogenetic analyses confirm the classification of D. ocellatus within Diploschistes. Three different groups have been recognized within this genus, based on ascomatal morphology: Actinostomus (perithecioid), Scruposus (urceolate), and Ocel- latus (lecanoroid). These groups have been widely used in monographic studies and keys, but their taxonomic value has not been confirmed yet. Here we inferred phylogenetic relationships within Diploschistes, with a special emphasis on the D. scruposus complex, using a combined dataset consisting of morphological, chemical, nrITS, and mtSSU data in order to determine if these species groups and phenotypically based species delimitations were monophyletic. Based on our results, a new subgeneric treatment for Diploschistes is proposed, and the taxonomic value of fruiting body types is confirmed. The clade correspond- ing to D. ocellatus consists of two well-supported subclades, one of them grouping specimens without ascomata, having only pycnidia. It is also remarkable that the clade containing specimens of D. diacapsis subsp. neutrophilus appears distantly related to the clade containing all other accessions of D. diacapsis. Our analysis revealed that for some taxa, such as D. scruposus and D. interpediens, molecular variability did not correlate with either morphological or chemical diversity.

First report of the pantropical species Diploschistes rampoddensis from Europe.

The lichen species Diploschistes rampoddensis, previously known only from a few localities in tropical and subtropical Asia and Oceania, is reported here for the first time in Europe. A detailed description, including macroand microscopic characters, and comparisons with closely related taxa are also provided. Molecular analyses based on the nrITS were used to confirm this new record for the European lichen biota.