Sergio Pérez-Ortega

Revisiting photobiont diversity in the lichen family Verrucariaceae (Ascomycota)

The Verrucariaceae (Ascomycota) is a family of mostly lichenized fungi with a unique diversity of algal symbionts, including some algae that are rarely or never associated with other lichens. The phylogenetic position of most of these algae has not yet been studied and, because morphology-based identifications can often be misleading, molecular data is necessary to revisit their identity and to explore patterns of association between fungal and algal partners. For this reason, the diversity of photobionts in this lichen family was investigated using molecular markers (rbcL and nuSSU) amplified from DNA extracts of lichen thalli and cultured isolates. Although a single algal genus, Diplosphaera (Trebouxiophyceae), was associated with 12 out of the 17 sampled genera of Verrucariaceae, representatives of eight other genera in five orders of the Chlorophyta and one genus in the Xanthophyceae also form lichen associations with members of the family. Fungal genera with simple crustose thalli (e.g. Hydropunctaria, Wahlenbergiella, Bagliettoa) use a high diversity and unusual selection of photobionts. In contrast, fungal genera with more complex thalli (e.g. Placidium, Dermatocarpon) tend to have lower photobiont diversity. Habitat requirements and phylogenetic histories are both partly reflected in the observed patterns of associations between lichenized fungi from the family Verrucariaceae and their photobionts.

Lichens and lichenicolous fungi of the Klondike Gold Rush National Historic Park, Alaska, in a global biodiversity context

Abstract The lichen flora of southeast Alaska has been explored for over 100 years, but remains poorly known. In the first survey of its kind from the region, we report 766 taxa of lichens and lichenicolous fungi from the Klondike Gold Rush National Historic Park. Coming from a park only 53 km2 in size, this represents one of the largest numbers of lichenized and lichenicolous fungi per unit area ever reported and the largest number ever reported from any United States National Park of any size. One lichen genus, four lichen species and one species of lichenicolous fungus are described as new to science: Steineropsis alaskana gen. et sp. nov. (Peltigerales), Coccotrema hahriae and Pertusaria mccroryae (both Pertusariales), Stereocaulon klondikense (Lecanorales) and Corticifraga scrobiculatae (Lecanoromycetes incertae sedis). The new combinations Coccotrema minutum and Nesolechia fusca are made and Parmelia elongata is reduced to synonymy under Hypogymnia duplicata. Seventy-five taxa could not be assigned a species name at this time and represent a pool of critical and/or potentially undescribed taxa. Thirty-four taxa are new or confirmed for North America, including seventeen lichen taxa, one species of saprophytic Dothideomycetes, and sixteen species of lichenicolous fungi. Five taxa of eastern North American distribution are reported for the west for the first time. A total of 196 taxa are new for Alaska. We report the presence of novel secondary chemical compounds in Bryoria, Cladonia, Hypogymnia and Pertusaria. Based on a Chao analysis of single and two-time occurrences we estimate we have captured not more than 83% of the macrolichen and 64% of the microlichen flora, with the total flora likely exceeding 1000 taxa. We provide an overview of the lichen inventories with highest species number worldwide and discuss the Klondike in the context of broader patterns in lichen richness. Global models of climate change in the coming century are unanimous in predicting greater temperature increases at higher latitudes than at low latitudes. We hope that our data will lead to a reconsideration of the potential extent of biodiversity at high latitudes.

Evolution of complex symbiotic relationships in a morphologically derived family of lichen-forming fungi.

We studied the evolutionary history of the Parmeliaceae (Lecanoromycetes, Ascomycota), one of the largest families of lichen-forming fungi with complex and variable morphologies, also including several lichenicolous fungi. We assembled a six-locus data set including nuclear, mitochondrial and low-copy protein-coding genes from 293 operational taxonomic units (OTUs). The lichenicolous lifestyle originated independently three times in lichenized ancestors within Parmeliaceae, and a new generic name is introduced for one of these fungi. In all cases, the independent origins occurred c. 24 million yr ago. Further, we show that the Paleocene, Eocene and Oligocene were key periods when diversification of major lineages within Parmeliaceae occurred, with subsequent radiations occurring primarily during the Oligocene and Miocene. Our phylogenetic hypothesis supports the independent origin of lichenicolous fungi associated with climatic shifts at the Oligocene-Miocene boundary. Moreover, diversification bursts at different times may be crucial factors driving the diversification of Parmeliaceae. Additionally, our study provides novel insight into evolutionary relationships in this large and diverse family of lichen-forming ascomycetes.

Photobiont selectivity leads to ecological tolerance and evolutionary divergence in a polymorphic complex of lichenized fungi

BACKGROUND AND AIMS The integrity and evolution of lichen symbioses depend on a fine-tuned combination of algal and fungal genotypes. Geographically widespread species complexes of lichenized fungi can occur in habitats with slightly varying ecological conditions, and it remains unclear how this variation correlates with symbiont selectivity patterns in lichens. In an attempt to address this question, >300 samples were taken of the globally distributed and ecologically variable lichen-forming species complex Tephromela atra, together with closely allied species, in order to study genetic diversity and the selectivity patterns of their photobionts. METHODS Lichen thalli of T. atra and of closely related species T. grumosa, T. nashii and T. atrocaesia were collected from six continents, across 24 countries and 62 localities representing a wide range of habitats. Analyses of genetic diversity and phylogenetic relationships were carried out both for photobionts amplified directly from the lichen thalli and from those isolated in axenic cultures. Morphological and anatomical traits were studied with light and transmission electron microscopy in the isolated algal strains. KEY RESULTS Tephromela fungal species were found to associate with 12 lineages of Trebouxia. Five new clades demonstrate the still-unrecognized genetic diversity of lichen algae. Culturable, undescribed lineages were also characterized by phenotypic traits. Strong selectivity of the mycobionts for the photobionts was observed in six monophyletic Tephromela clades. Seven Trebouxia lineages were detected in the poorly resolved lineage T. atra sensu lato, where co-occurrence of multiple photobiont lineages in single thalli was repeatedly observed. CONCLUSIONS Low selectivity apparently allows widespread lichen-forming fungi to establish successful symbioses with locally adapted photobionts in a broader range of habitats. This flexibility might correlate with both lower phylogenetic resolution and evolutionary divergence in species complexes of crustose lichen-forming fungi.

Lichen myco- and photobiont diversity and their relationships at the edge of life (McMurdo Dry Valleys, Antarctica)

Lichen-forming fungi are among the most diverse group of organisms in Antarctica. Being poikilohydric, lichens are able to cope with harsh environmental conditions that exclude other organisms like vascular plants. The McMurdo Dry Valleys (Victoria Land, Continental Antarctica) are a hyperarid cold desert where macroscopic life is reduced to a few lichens and bryophyte species. We investigated the diversity of lichen-forming fungi and their associated photobionts in three valleys (Garwood, Marshall, and Miers). Correct identification of lichen-forming fungi from extreme ecosystems is complicated by the presence of numerous sterile and extremely modified thalli. To overcome this problem, we used a combined approach for the identification of the species present in the area, the first involving identification by means of standard characters and the second, two DNA-based (ITS region) species delimitation methods (General Mixed Yule-Coalescent model and genetic distances). In addition, we also used ITS sequences for the identification of the photobionts associated with the mycobionts. We studied the relationships between both bionts and assessed the degree of selectivity and specificity found in those associations. We also looked for landscape level spatial patterns in these associations. The two DNA-based methods performed quite differently, but 27 species of lichen-forming fungi and five putative species of photobionts were found in the studied area. Although there was a general trend for low selectivity in the relationships, high specificity was found in some associations and differential selectivity was observed in some lichen-forming fungi. No spatial structure was detected in the distribution of photobionts in the studied area.

Symbiotic lifestyle and phylogenetic relationships of the bionts of Mastodia tessellata (Ascomycota, incertae sedis).

The biological nature of some symbioses is unclear because it is often not easy to discern whether the symbionts obtain any benefits from the association. Mastodia tessellata, a symbiosis between a leafy green alga and a fungus of uncertain phylogenetic position, is among the most investigated, controversial, and poorly understood associations. Because it has been difficult to determine whether this association is mutually beneficial or parasitic, not all scientists accept M. tessellata as a true lichen symbiosis. Mastodia tessellata is thus an interesting model to illustrate the interactions and processes that occur in fungal-algal symbioses. To improve our understanding of this association, we address the phylogenetic positions of the bionts involved and examine their interactions at the ultrastructural level. Examining the nuLSU and nuSSU gene regions of the mycobiont and the rbcL gene region of the photobiont, we found the fungus to be related to a group of marine species in the genus Verrucaria, family Verrucariaceae, despite its present ascription to the family Mastodiaceae. In addition, the photobiont of the symbiosis emerged as closely related to the North American species Prasiola borealis. Our electron microscopy observations provide new information on the process of fungal colonization of the algal thalli, as well as on relationships between the symbionts during different stages of colonization. The special features of this lichen symbiosis are discussed and compared with other examples of fungal symbioses in nature.

Global assessment of genetic variation and phenotypic plasticity in the lichen-forming species Tephromela atra

Understanding how many species exist and the processes by which they form remains a central topic of ecological and evolutionary biology, but represents a special challenge within microbial groups. The lichen-forming fungi represent one of the best examples in which species evolution and diversity create patterns of high phenotypic plasticity coupled with wide geographic distributions. We sampled the lichen-forming species Tephromela atra and related species at a world-wide scale to reconstruct a phylogenetic hypothesis using three nuclear markers. Samples were also studied for morphological and chemical traits to assess how well the phenotypic relationships with species, previously segregated from T. atra, agrees with molecular data. We used a genealogical concordance approach and identified 15 monophyletic clades, which may represent independent lineages. By combining morphological and chemical characters, ecological preferences and geographic origin we distinguish six different species. Although subtle phenotypical traits are frequently used for describing previously cryptic species in fungi, the continuum of variability found in morphology and chemical patterns in T. atra prevents the description of new taxa with characteristic traits. We observed that phenotypic characters arise in parallel at local or regional scale but are not correlated with genetic isolation. Therefore, they are insufficient for characterizing species with broad geographic ranges within T. atra.

Austroparmelina, a new Australasian lineage in parmelioid lichens (Parmeliaceae, Ascomycota)

Parmelioid lichens form the largest monophyletic group within the Parmeliaceae, a family distributed worldwide. The genus Parmelina was described by Hale (1976a) accommodating species from both hemispheres. We have employed parsimony, Bayesian and maximum likelihood analyses of a combined data set of nu ITS, LSU and mt SSU rDNA sequences to (1) test the monophyly of Parmelina and (2) to elucidate the generic status and phylogenetic position of the Australasian species. Twenty-one new sequences were generated in this study. Our results provide evidence that Parmelina is polyphyletic and the species fall into two major well-supported groups (Groups I and II). The Australasian species of Parmelina and two species of Canoparmelia (C. pruinata and C. macrospora) form Group I, which is nested within the parmotremoid genera of Parmeliaceae, Parmelina species from the northern hemisphere including those from western North America and the Mediterranean basin form a monophyletic group (Group II), which is sister to the East Asian temperate genus Myelochroa. Morphological and chemical features were reevaluated considering this observed phylogeny. Some morphological features like lobe morphology, several traits in the excipulum and geography are useful in characterizing the monophyletic lineage of the Australasian Parmelina/Canoparmelina species. This lineage is described as the new genus Austroparmelina. Thirteen new combinations in the new genus are proposed.

A molecular phylogeny of the Lecanora varia group, including a new species from western North America

The phylogeny of the Lecanora varia group is inferred from ITS sequences using Bayesian and ML phylogenetic analysis methods. According to our data, usnic acid-containing, corticolous Lecanora species do not form a monophyletic group but occur in three major clades together with other groups of Lecanora and Protoparmeliopsis. The new combination Lecanora filamentosa (Stirt.) Elix & Palice is proposed. The new species Lecanora schizochromatica is described from northwestern North America. It produces atranorin as a major substance and is closely related to the morphologically and chemically similar L. filamentosa. The American Biatora pullula Tuck. is synonymised with Lecanora anopta Nyl., which is hereby reported for continental North America for the first time. The phylogenetic relationships between the major clades of Lecanora are still largely unresolved and require more intensive taxon and character sampling.

Extreme phenotypic variation in Cetraria aculeata (lichenized Ascomycota): adaptation or incidental modification?

BACKGROUND AND AIMS Phenotypic variability is a successful strategy in lichens for colonizing different habitats. Vagrancy has been reported as a specific adaptation for lichens living in steppe habitats around the world. Among the facultatively vagrant species, the cosmopolitan Cetraria aculeata apparently forms extremely modified vagrant thalli in steppe habitats of Central Spain. The aim of this study was to investigate whether these changes are phenotypic plasticity (a single genotype producing different phenotypes), by characterizing the anatomical and ultrastructural changes observed in vagrant morphs, and measuring differences in ecophysiological performance. METHODS Specimens of vagrant and attached populations of C. aculeata were collected on the steppes of Central Spain. The fungal internal transcribed spacer (ITS), glyceraldehyde-3-phosphate dehydrogenase (GPD) and the large sub-unit of the mitochondrial ribosomal DNA (mtLSUm), and the algal ITS and actin were studied within a population genetics framework. Semi-thin and ultrathin sections were analysed by means of optical, scanning electron and transmission electron microscopy. Gas exchange and chlorophyll fluorescence were used to compare the physiological performance of both morphs. KEY RESULTS AND CONCLUSIONS Vagrant and attached morphs share multilocus haplotypes which may indicate that they belong to the same species in spite of their completely different anatomy. However, differentiation tests suggested that vagrant specimens do not represent a random sub-set of the surrounding population. The morphological differences were related to anatomical and ultrastructural differences. Large intercalary growth rates of thalli after the loss of the basal-apical thallus polarity may be the cause of the increased growth shown by vagrant specimens. The anatomical and morphological changes lead to greater duration of ecophysiological activity in vagrant specimens. Although the anatomical and physiological changes could be chance effects, the genetic differentiation between vagrant and attached sub-populations and the higher biomass of the former show fitness effects and adaptation to dry environmental conditions in steppe habitats.