Circadian clock- and temperature-associated genes contribute to overall genomic differentiation along elevation in lichenized fungi.

Circadian regulation is linked to local environmental adaptation, and many species with broad climatic niches display variation in circadian genes. Here, we hypothesize that lichenizing fungi occupying different climate zones tune their metabolism to local environmental conditions with the help of their circadian systems. We study two species of the genus Umbilicaria occupying similar climatic niches (Mediterranean and the cold temperate) in different continents. Using homology to Neurospora crassa genes, we identify gene sets associated with circadian rhythms (11 core, 39 peripheral genes) as well as temperature response (37 genes). Nucleotide diversity of these genes is significantly correlated with mean annual temperature, minimum temperature of the coldest month and mean temperature of the coldest quarter. Furthermore, we identify altitudinal clines in allele frequencies in several non-synonymous substitutions in core clock components, for example, white collar-like, frh-like and various ccg-like genes. A dN/dS approach revealed a few significant peripheral clock- and temperature-associated genes (e.g. ras-1-like, gna-1-like) that may play a role in fine-tuning the circadian clock and temperature-response machinery. An analysis of allele frequency changes demonstrated the strongest evidence for differentiation above the genomic background in the clock-associated genes in U. pustulata. These results highlight the likely relevance of the circadian clock in environmental adaptation, particularly frost tolerance, of lichens. Whether or not the fungal clock modulates the symbiotic interaction within the lichen consortium remains to be investigated. We corroborate the finding of genetic variation in clock components along altitude-not only latitude-as has been reported in other species.

The 4D virtual patient: A proof of concept in digital dentistry.

This dental technique article updates the 3-dimensional (3D) virtual patient digital workflow by introducing the space-time information acquired with jaw tracking motion. The direct digital procedure involved scanning the interim prostheses, creating virtual casts, and transferring the digital facebow, virtual articulation, and jaw-tracking motion. All 3D files and records from the intraoral scanner, cone beam computed tomography, extraoral scanner, and jaw tracking motion were superimposed using existing teeth as the connecting link in all data sets and adopting the best-fit alignment. The main advantages of the technique were creating a 4-dimensional virtual patient and standardizing the introduction of clinical space-time information (the fourth dimension) beyond the static condition.

Maxillomandibular relationship and virtual facebow integration in complete-arch intraoral implant scan: A novel clinical technique.

This clinical report introduces a novel clinical technique to create a 3D virtual patient for transferring the edentulous maxillary arch position with maxillomandibular relationship by using a facial scan device and an intraoral scanner and omitting CBCT imaging.

Lichen holobionts show compositional structure along elevation.

Holobionts are dynamic ecosystems that may respond to abiotic drivers with compositional changes. Uncovering elevational diversity patterns within these microecosystems can further our understanding of community-environment interactions. Here, we assess how the major components of lichen holobionts-fungal hosts, green algal symbionts, and the bacterial community-collectively respond to an elevational gradient. We analyse populations of two lichen symbioses, Umbilicaria pustulata and U. hispanica, along an elevational gradient spanning 2100 altitudinal metres and covering three major biomes. Our study shows (i) discontinuous genomic variation in fungal hosts with one abrupt genomic differentiation within each of the two host species, (ii) altitudinally structured bacterial communities with pronounced turnover within and between hosts, and (iii) altitude-specific presence of algal symbionts. Alpha diversity of bacterial communities decreased with increasing elevation. A marked turnover in holobiont diversity occurred across two altitudinal belts: at 11°C-13°C average annual temperature (here: 800-1200 m a.s.l.), and at 7°C-9°C average annual temperature (here: 1500-1800 m a.s.l.). The two observed zones mark a clustering of distribution limits and community shifts. The three ensuing altitudinal classes, that is, the most frequent combinations of species in holobionts, approximately correspond to the Mediterranean, cool-temperate, and alpine climate zones. We conclude that multitrophic microecosystems, such as lichen holobionts, respond with concerted compositional changes to climatic factors that also structure communities of macroorganisms, for example, vascular plants.

Metatranscriptomics reveals contrasting effects of elevation on the activity of bacteria and bacterial viruses in soil.

Soil microbial diversity affects ecosystem functioning and global biogeochemical cycles. Soil bacterial communities catalyse a diversity of biogeochemical reactions and have thus sparked considerable scientific interest. One driver of bacterial community dynamics in natural ecosystems has so far been largely neglected: the predator-prey interactions between bacterial viruses (bacteriophages) and bacteria. To generate ground level knowledge on environmental drivers of these particular predator-prey dynamics, we propose an activity-based ecological framework to simultaneous capture community dynamics of bacteria and bacteriophages in soils. An ecological framework and specifically the analyses of community dynamics across latitudinal and elevational gradients have been widely used in ecology to understand community-wide responses of innumerable taxa to environmental change, in particular to climate. Here, we tested the hypothesis that the activity of bacteria and bacteriophages codeclines across an elevational gradient. We used metatranscriptomics to investigate bacterial and bacteriophage activity patterns at five sites across 400 elevational metres in the Swiss Alps in 2015 and 2017. We found that metabolic activity (transcription levels) of bacteria declined significantly with increasing elevation, but activity of bacteriophages did not. We showed that bacteriophages are consistently active in soil along the entire gradient, making bacteriophage activity patterns divergent from that of their putative bacterial prey. Future efforts will be necessary to link the environment-activity relationship to predator-prey dynamics, and to understand the magnitude of viral contributions to carbon, nitrogen and phosphorus cycling when infection causes bacterial cell death, a process that may represent an overlooked component of soil biogeochemical cycles.

Hygienic maintenance in patients with maxillae atrophy and in oncological patients with maxillary resection rehabilitated with zygomatic implants: A nested monocentric prospective cohort study.

OBJECTIVE: To assess peri-implant soft tissues condition, comparing clinical parameters of implants placed in patients with atrophic upper jaws and patients who underwent maxillary resection for oncological reasons. BACKGROUND: Zygomatic implants (ZIs) in oncologic patients could be affected by more complications compared to implants placed in atrophic maxillae. The soft tissue condition is an essential requirement for implant success, but few studies have investigated this topic. METHODS: A nested monocentric prospective parallel cohort (atrophic vs. oncological patients) study was performed. Clinical visits and professional hygiene sessions were performed every three months, and bleeding on probing (BOP), probing pocket depth (PPD), gingival index (GI), plaque index (PI) and implant mobility were recorded by a blind outcome assessor. RESULTS: In total, 77 ZIs placed in 21 patients were evaluated: 54 (70.1%) ZIs were inserted in patients belonging to the atrophic cohort (PAM) and 23 (29.9%) ZIs in the oncologic cohort (OP). The probability of having BOP at the considered mean follow-up (27 months) was 24.8% (95% CI 19.0-31.9) for PAM and 22.9% (95% CI 15.1-33.9) OP. The mean PPD values were 2.78 ± 1.28 (range 1-8) in PAM and 2.91 ± 1.98 (range 0-10) in OP. None of the implants showed mobility. No associations between group belongingness and the entity of PPD, PI, GI and the risk of BOP were found, adjusting for the considered confounding factors (age, smoking and implant position). CONCLUSIONS: Under a strict supportive hygiene therapy protocol ZIs in oncologic patients showed similar peri-implant tissue conditions to that of patients with maxillary atrophy.

Virus diversity in metagenomes of a lichen symbiosis (Umbilicaria phaea): complete viral genomes, putative hosts and elevational distributions.

Viruses can play critical roles in symbioses by initiating horizontal gene transfer, affecting host phenotypes, or expanding their host's ecological niche. However, knowledge of viral diversity and distribution in symbiotic organisms remains elusive. Here we use deep-sequenced metagenomic DNA (PacBio Sequel II; two individuals), paired with a population genomics approach (Pool-seq; 11 populations, 550 individuals) to understand viral distributions in the lichen Umbilicaria phaea. We assess (i) viral diversity in lichen thalli, (ii) putative viral hosts (fungi, algae, bacteria) and (iii) viral distributions along two replicated elevation gradients. We identified five novel viruses, showing 28%-40% amino acid identity to known viruses. They tentatively belong to the families Caulimoviridae, Myoviridae, Podoviridae and Siphoviridae. Our analysis suggests that the Caulimovirus is associated with green algal photobionts (Trebouxia) of the lichen, and the remaining viruses with bacterial hosts. We did not detect viral sequences in the mycobiont. Caulimovirus abundance decreased with increasing elevation, a pattern reflected by a specific algal lineage hosting this virus. Bacteriophages showed population-specific patterns. Our work provides the first comprehensive insights into viruses associated with a lichen holobiont and suggests an interplay of viral hosts and environment in structuring viral distributions.

Climate-specific biosynthetic gene clusters in populations of a lichen-forming fungus.

Natural products can contribute to abiotic stress tolerance in plants and fungi. We hypothesize that biosynthetic gene clusters (BGCs), the genomic elements that underlie natural product biosynthesis, display structured differences along elevation gradients. We analysed biosynthetic gene variation in natural populations of the lichen-forming fungus Umbilicaria pustulata. We collected a total of 600 individuals from the Mediterranean and cold-temperate climates. Population genomic analyses indicate that U. pustulata contains three clusters that are highly differentiated between the Mediterranean and cold-temperate populations. One entire cluster is exclusively present in cold-temperate populations, and a second cluster is putatively dysfunctional in all cold-temperate populations. In the third cluster variation is fixed in all cold-temperate populations due to hitchhiking. In these two clusters the presence of consistent allele frequency differences among replicate populations/gradients suggests that selection rather than drift is driving the pattern. We advocate that the landscape of fungal biosynthetic genes is shaped by both positive and hitchhiking selection. We demonstrate, for the first time, the presence of climate-associated BGCs and BGC variations in lichen-forming fungi. While the associated secondary metabolites of the candidate clusters are presently unknown, our study paves the way for targeted discovery of natural products with ecological significance.

Expanding the mutualistic niche: parallel symbiont turnover along climatic gradients.

Keystone mutualisms, such as corals, lichens or mycorrhizae, sustain fundamental ecosystem functions. Range dynamics of these symbioses are, however, inherently difficult to predict because host species may switch between different symbiont partners in different environments, thereby altering the range of the mutualism as a functional unit. Biogeographic models of mutualisms thus have to consider both the ecological amplitudes of various symbiont partners and the abiotic conditions that trigger symbiont replacement. To address this challenge, we here investigate 'symbiont turnover zones'--defined as demarcated regions where symbiont replacement is most likely to occur, as indicated by overlapping abundances of symbiont ecotypes. Mapping the distribution of algal symbionts from two species of lichen-forming fungi along four independent altitudinal gradients, we detected an abrupt and consistent ß-diversity turnover suggesting parallel niche partitioning. Modelling contrasting environmental response functions obtained from latitudinal distributions of algal ecotypes consistently predicted a confined altitudinal turnover zone. In all gradients this symbiont turnover zone is characterized by approximately 12°C average annual temperature and approximately 5°C mean temperature of the coldest quarter, marking the transition from Mediterranean to cool temperate bioregions. Integrating the conditions of symbiont turnover into biogeographic models of mutualisms is an important step towards a comprehensive understanding of biodiversity dynamics under ongoing environmental change.

The lichen symbiosis re-viewed through the genomes of Cladonia grayi and its algal partner Asterochloris glomerata.

BACKGROUND: Lichens, encompassing 20,000 known species, are symbioses between specialized fungi (mycobionts), mostly ascomycetes, and unicellular green algae or cyanobacteria (photobionts). Here we describe the first parallel genomic analysis of the mycobiont Cladonia grayi and of its green algal photobiont Asterochloris glomerata. We focus on genes/predicted proteins of potential symbiotic significance, sought by surveying proteins differentially activated during early stages of mycobiont and photobiont interaction in coculture, expanded or contracted protein families, and proteins with differential rates of evolution. RESULTS: A) In coculture, the fungus upregulated small secreted proteins, membrane transport proteins, signal transduction components, extracellular hydrolases and, notably, a ribitol transporter and an ammonium transporter, and the alga activated DNA metabolism, signal transduction, and expression of flagellar components. B) Expanded fungal protein families include heterokaryon incompatibility proteins, polyketide synthases, and a unique set of G-protein α subunit paralogs. Expanded algal protein families include carbohydrate active enzymes and a specific subclass of cytoplasmic carbonic anhydrases. The alga also appears to have acquired by horizontal gene transfer from prokaryotes novel archaeal ATPases and Desiccation-Related Proteins. Expanded in both symbionts are signal transduction components, ankyrin domain proteins and transcription factors involved in chromatin remodeling and stress responses. The fungal transportome is contracted, as are algal nitrate assimilation genes. C) In the mycobiont, slow-evolving proteins were enriched for components involved in protein translation, translocation and sorting. CONCLUSIONS: The surveyed genes affect stress resistance, signaling, genome reprogramming, nutritional and structural interactions. The alga carries many genes likely transferred horizontally through viruses, yet we found no evidence of inter-symbiont gene transfer. The presence in the photobiont of meiosis-specific genes supports the notion that sexual reproduction occurs in Asterochloris while they are free-living, a phenomenon with implications for the adaptability of lichens and the persistent autonomy of the symbionts. The diversity of the genes affecting the symbiosis suggests that lichens evolved by accretion of many scattered regulatory and structural changes rather than through introduction of a few key innovations. This predicts that paths to lichenization were variable in different phyla, which is consistent with the emerging consensus that ascolichens could have had a few independent origins.

Erysipelas vaccination protocols in dolphins Tursiops truncatus evaluated by antibody responses over twenty continuous years.

Erysipelas is an infection caused by Erysipelothrix rhusiopathiae that affects many different species around the world, including cetaceans. The acute septicemic form can rapidly cause death in bottlenose dolphins Tursiops truncatus. The ultimate goals of this long-term study were the development and identification of the most effective vaccination protocol against clinical erysipelas in T. truncatus using a commercially available swine vaccine, and to determine whether there is a need for a semi-annual vaccination versus an annual vaccination. The present study concentrated on the immunization of a dolphin population (7 wild-born and 22 captive-born individuals) with 2 swine vaccines, the European 'Eurovac Ery®' vaccine and the American 'ER Bac Plus®' vaccine, and immunological profile results over a 20-yr time period. The general protocol was a primo-vaccination (between 3 and 7 mo of age for calves) with or without a booster 1 mo post primo-vaccination and either annual or semi-annual vaccination thereafter. Sera were collected prior to vaccination, 2 wk post-vaccination and monthly. A dolphin-specific ELISA was developed to analyze the erysipelas-specific antibody response of vaccinated animals. The final ELISA results (n = 1362 samples from 29 animals at pre- and post-vaccination time) suggest that (1) there is a significant difference in antibody levels at the start of the vaccination between older and younger animals; (2) at least 3 vaccinations are necessary to obtain antibody levels above the levels at pre-vaccination; (3) thereafter, annual vaccinations seem sufficient to keep antibody levels above the levels at pre-vaccination; and (4) both vaccines induced similar responses. No case of erysipelas infection was observed in this population during the study.

Biosynthetic Gene Content of the 'Perfume Lichens' Evernia prunastri and Pseudevernia furfuracea.

Lichen-forming fungi produce a vast number of unique natural products with a wide variety of biological activities and human uses. Although lichens have remarkable potential in natural product research and industry, the molecular mechanisms underlying the biosynthesis of lichen metabolites are poorly understood. Here we use genome mining and comparative genomics to assess biosynthetic gene clusters and their putative regulators in the genomes of two lichen-forming fungi, which have substantial commercial value in the perfume industry, Evernia prunastri and Pseudevernia furfuracea. We report a total of 80 biosynthetic gene clusters (polyketide synthases (PKS), non-ribosomal peptide synthetases and terpene synthases) in E. prunastri and 51 in P. furfuracea. We present an in-depth comparison of 11 clusters, which show high homology between the two species. A ketosynthase (KS) phylogeny shows that biosynthetic gene clusters from E. prunastri and P. furfuracea are widespread across the Fungi. The phylogeny includes 15 genomes of lichenized fungi and all fungal PKSs with known functions from the MIBiG database. Phylogenetically closely related KS domains predict not only similar PKS architecture but also similar cluster architecture. Our study highlights the untapped biosynthetic richness of lichen-forming fungi, provides new insights into lichen biosynthetic pathways and facilitates heterologous expression of lichen biosynthetic gene clusters.

Environment and host identity structure communities of green algal symbionts in lichens.

An understanding of how biotic interactions shape species' distributions is central to predicting host-symbiont responses under climate change. Switches to locally adapted algae have been proposed to be an adaptive strategy of lichen-forming fungi to cope with environmental change. However, it is unclear how lichen photobionts respond to environmental gradients, and whether they play a role in determining the fungal host's upper and lower elevational limits. Deep-coverage Illumina DNA metabarcoding was used to track changes in the community composition of Trebouxia algae associated with two phylogenetically closely related, but ecologically divergent fungal hosts along a steep altitudinal gradient in the Mediterranean region. We detected the presence of multiple Trebouxia species in the majority of thalli. Both altitude and host genetic identity were strong predictors of photobiont community assembly in these two species. The predominantly clonally dispersing fungus showed stronger altitudinal structuring of photobiont communities than the sexually reproducing host. Elevation ranges of the host were not limited by the lack of compatible photobionts. Our study sheds light on the processes guiding the formation and distribution of specific fungal-algal combinations in the lichen symbiosis. The effect of environmental filtering acting on both symbiotic partners appears to shape the distribution of lichens.

Antimicrobial-producing Pseudoalteromonas from the marine environment of Panama shows a high phylogenetic diversity and clonal structure.

Pseudoalteromonas is a genus of marine bacteria often found in association with other organisms. Although several studies have examined Pseudoalteromonas diversity and their antimicrobial activity, its diversity in tropical environments is largely unexplored. We investigated the diversity of Pseudoalteromonas in marine environments of Panama using a multilocus phylogenetic approach. Furthermore we tested their antimicrobial capacity and evaluated the effect of recombination and mutation in shaping their phylogenetic relationships. The reconstruction of clonal relationships among 78 strains including 15 reference Pseudoalteromonas species revealed 43 clonal lineages, divided in pigmented and non-pigmented strains. In total, 39 strains displayed moderate to high activity against Gram-positive and Gram-negative bacteria and fungi. Linkage disequilibrium analyses showed that the Pseudoalteromonas strains of Panama have a highly clonal structure and that, although present, recombination is not frequent enough to break the association among alleles. This clonal structure is in contrast to the high rates of recombination generally reported for aquatic and marine bacteria. We propose that this structure is likely due to the symbiotic association with marine invertebrates of most strains analyzed. Our results also show that there are several putative new species of Pseudoalteromonas in Panama to be described.

Adaptive differentiation coincides with local bioclimatic conditions along an elevational cline in populations of a lichen-forming fungus.

BACKGROUND: Many fungal species occur across a variety of habitats. Particularly lichens, fungi forming symbioses with photosynthetic partners, have evolved remarkable tolerances for environmental extremes. Despite their ecological importance and ubiquity, little is known about the genetic basis of adaption in lichen populations. Here we studied patterns of genome-wide differentiation in the lichen-forming fungus Lasallia pustulata along an altitudinal gradient in the Mediterranean region. We resequenced six populations as pools and identified highly differentiated genomic regions. We then detected gene-environment correlations while controlling for shared population history and pooled sequencing bias, and performed ecophysiological experiments to assess fitness differences of individuals from different environments. RESULTS: We detected two strongly differentiated genetic clusters linked to Mediterranean and temperate-oceanic climate, and an admixture zone, which coincided with the transition between the two bioclimates. High altitude individuals showed ecophysiological adaptations to wetter and more shaded conditions. Highly differentiated genome regions contained a number of genes associated with stress response, local environmental adaptation, and sexual reproduction. CONCLUSIONS: Taken together our results provide evidence for a complex interplay between demographic history and spatially varying selection acting on a number of key biological processes, suggesting a scenario of ecological speciation.

Fungal-algal association patterns in lichen symbiosis linked to macroclimate.

Both macroclimate and evolutionary events may influence symbiont association and diversity patterns. Here we assess how climatic factors and evolutionary events shape fungal-algal association patterns in the widely distributed lichen-forming fungal genus Protoparmelia. Multilocus phylogenies of fungal and algal partners were generated using 174 specimens. Coalescent-based species delimitation analysis suggested that 23 fungal hosts are associating with 20 algal species. Principal component analysis (PCA) was performed to infer how fungal-algal association patterns varied with climate. Fungi associated with one to three algal partners whereas algae accepted one to five fungal partners. Both fungi and algae were more specific, associating with fewer partners, in the warmer climates. Interaction with more than one partner was more frequent in cooler climates for both the partners. Cophylogenetic analyses suggest congruent fungal-algal phylogenies. Host switch was a more common event in warm climates, whereas failure of the photobiont to diverge with its fungal host was more frequent in cooler climates. We conclude that both environmental factors and evolutionary events drive fungal and algal evolution in Protoparmelia. The processes leading to phylogenetic congruence of fungi and algae are different in different macrohabitats in our study system. Hence, closely related species inhabiting diverse habitats may follow different evolutionary pathways.

Genomic and expression analyses of Tursiops truncatus T cell receptor gamma (TRG) and alpha/delta (TRA/TRD) loci reveal a similar basic public γδ repertoire in dolphin and human.

BACKGROUND: The bottlenose dolphin (Tursiops truncatus) is a mammal that belongs to the Cetartiodactyla and have lived in marine ecosystems for nearly 60 millions years. Despite its popularity, our knowledge about its adaptive immunity and evolution is very limited. Furthermore, nothing is known about the genomics and evolution of dolphin antigen receptor immunity. RESULTS: Here we report a evolutionary and expression study of Tursiops truncatus T cell receptor gamma (TRG) and alpha/delta (TRA/TRD) genes. We have identified in silico the TRG and TRA/TRD genes and analyzed the relevant mature transcripts in blood and in skin from four subjects. The dolphin TRG locus is the smallest and simplest of all mammalian loci as yet studied. It shows a genomic organization comprising two variable (V1 and V2), three joining (J1, J2 and J3) and a single constant (C), genes. Despite the fragmented nature of the genome assemblies, we deduced the TRA/TRD locus organization, with the recent TRDV1 subgroup genes duplications, as it is expected in artiodactyls. Expression analysis from blood of a subject allowed us to assign unambiguously eight TRAV genes to those annotated in the genomic sequence and to twelve new genes, belonging to five different subgroups. All transcripts were productive and no relevant biases towards TRAV-J rearrangements are observed. Blood and skin from four unrelated subjects expression data provide evidence for an unusual ratio of productive/unproductive transcripts which arise from the TRG V-J gene rearrangement and for a "public" gamma delta TR repertoire. The productive cDNA sequences, shared both in the same and in different individuals, include biases of the TRGV1 and TRGJ2 genes. The high frequency of TRGV1-J2/TRDV1- D1-J4 productive rearrangements in dolphins may represent an interesting oligo-clonal population comparable to that found in human with the TRGV9- JP/TRDV2-D-J T cells and in primates. CONCLUSIONS: Although the features of the TRG and TRA/TRD loci organization reflect those of the so far examined artiodactyls, genomic results highlight in dolphin an unusually simple TRG locus. The cDNA analysis reveal productive TRA/TRD transcripts and unusual ratios of productive/unproductive TRG transcripts. Comparing multiple different individuals, evidence is found for a "public" gamma delta TCR repertoire thus suggesting that in dolphins as in human the gamma delta TCR repertoire is accompanied by selection for public gamma chain.

Taxonomic revision and species delimitation of coccoid green algae currently assigned to the genus Dictyochloropsis (Trebouxiophyceae, Chlorophyta).

Coccoid green algae traditionally classified in Dictyochloropsis have a complex, reticulate chloroplast, when mature, without a pyrenoid. They occupy remarkably diverse ecological niches as free-living organisms or in association with lichen-forming fungi and were recently shown to form two distinct lineages within Trebouxiophyceae. We used a polyphasic approach to revise the taxonomy of the genus. Based on phylogenetic analysis of the 18S rRNA gene, and detailed morphological investigation using comparative conventional light and confocal microscopy, we have assigned these lineages to two genera, Dictyochloropsis and Symbiochloris gen. nov. We have reconsidered the diagnostic generic features as follows: Dictyochloropsis comprises only free-living algae with a reticulate chloroplast, forming lobes in a parallel arrangement at some ontogenetic stages, and which reproduce only by means of autospores. This agrees with Geitler's original diagnosis of Dictyochloropsis, but not with the later emendation by Tschermak-Woess. Consequently, the species of Dictyochloropsis sensu Tschermak-Woess are assigned to Symbiochloris, with new combinations proposed. Symbiochloris encompasses free-living and/or lichenized algae with lobed chloroplasts and that reproduce by forming zoospores characterized by two subapical isokont flagella that emerge symmetrically near the flattened apex. In addition, using coalescent-based approaches, morphological characters and secondary structure of ITS transcripts, we inferred species boundaries and taxonomic relationships within the newly proposed genera. Two species of Dictyochloropsis and nine species of Symbiochloris are delimited, including the newly described species D. asterochloroides, S. handae, S. tropica, and S. tschermakiae. Our results further support the non-monophyly of autosporine taxa within Trebouxiophyceae.

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.

Molecular phylogeny and symbiotic selectivity of the green algal genus Dictyochloropsis s.l. (Trebouxiophyceae): a polyphyletic and widespread group forming photobiont-mediated guilds in the lichen family Lobariaceae.

Dictyochloropsis s.l. is an ecologically important, common but little-studied genus of green algae. Here, we examined the diversity and host selectivity of algae attributed to this genus at both species-to-species and species-to-community levels. We conducted a molecular investigation of 15 cultured strains and several lichen photobionts, using 18S rRNA, rbcL and ITS sequence data. We further used seven alga-specific microsatellite markers to study algal sharing among fungi of the family Lobariaceae in two populations in Madeira and Taiwan (454 lichens). We found that the genus Dictyochloropsis s.l. is polyphyletic. Dictyochloropsis clade 1 comprises only free-living algae whereas Dictyochloropsis clade 2 includes lichenized algae as well as free-living algae. Fungal selectivity towards algae belonging to Dictyochloropsis clade 2 is high. Selectivity varies geographically, with photobionts being restricted to a single region. Finally, we showed that Dictyochloropsis clade 2 individuals are shared among different fungal hosts in communities of lichens of the Lobariaceae. As for other green algal lineages, there is a high amount of cryptic diversity in Dictyochloropsis. Furthermore, co-evolution between Dictyochloropsis clade 2 algae and representatives of the Lobariaceae is manifested at the community level, with several unrelated fungal species being horizontally connected by shared photobiont clones.