Doctor of Philosophy / Department of Entomology / Ming-Shun Chen / John C. Reese / Keeping in view the important roles of bacteria in almost every aspect of insect’s life, the current study is the first systemic and intensive work on microbes associated with Hessian fly, a serious pest of wheat crop. A whole body analysis of Hessian fly larvae, pupae, or adults suggested that a remarkable diversity of bacteria is associated with different stages of the insect life cycle. The overriding detection of genera Acinetobacter and Enterobacter throughout the life cycle of Hessian fly suggested a stable and intimate relationship with the insect host. Adult Hessian flies have the most dissimilar bacterial composition from other stages with Bacillus as the most dominant genus. Analysis of 5778 high quality sequence reads obtained from larval gut estimated 187, 142, and 262 operational taxonomic units at 3% distance level from the 1st, 2nd, and 3rd instar respectively. Pseudomonas was the most dominant genus found in the gut of all three instars. The 3rd instar larval gut had the most diverse bacterial composition including genera Stenotrophomonas, Pantoea, Enterobacter, Ensifer, and Achromobacter. The transovarial transmission of major bacterial groups provided evidence of their intimate relationship with the Hessian fly. The Hessian fly is known to manipulate wheat plants to its own advantage. This study demonstrated that the combination of a decrease in carbon compounds and an increase in nitrogen compounds in the feeding tissues of Hessian fly-infested plants results in a C/N ratio of 17:1, nearly 2.5 times less than the C/N ratio (42:1) observed in control plants. We propose that bacteria associated with Hessian fly perform nitrogen fixation in the infested wheat, which was responsible for shifting the C/N ratio. The following findings made in the current study i.e. the presence of bacteria encoding nitrogenase (nifH) genes both in Hessian fly and infested wheat, exclusive expression of nifH in infested wheat, presence of diverse bacteria (including the nitrogen fixing genera) in the Hessian fly larvae, presence of similar bacterial microbiota in Hessian fly larvae and at the feeding site tissues in the infested wheat, and reduction in survival of Hessian fly larvae due to loss of bacteria are consistent with this hypothesis. The reduction in Hessian fly longevity after the loss of Alphaproteobacteria in first instar larvae, highest proportion of Alphaproteobacteria in insects surviving after the antibiotic treatments and the nitrogen fixation ability of associated Alphaproteobacteria strongly implies that Alphaproteobacteria are critical for the survival of Hessian fly larvae. This study provides a foundation for future studies to elucidate the role of associated microbes on Hessian fly virulence and biology. A better understanding of Hessian fly-microbe interactions may lead to new strategies to control this pest.

Vestimentiferan tubeworms of the eastern Pacific Ocean are often keystone species in vent communities. These polychaetes are host to intracellular Gammaproteobacteria symbionts. In this association, the siboglinid worms supply their symbionts with the compounds necessary to chemosynthesis while the sulfide oxidizing bacteria provide their host with the organic molecules necessary for their metabolism. The adult worms lack a digestive system and are therefore completely dependent on their symbionts for their nutrition. Given the obligate nature of the association for the host, it is surprising that the symbionts are not transmitted from parents to offspring but are acquired de novo from the environment at each generation. In other known cases of horizontally acquired mutualism (e.g. Rhizobium-legumes, dinoflagellates- corals), obtaining symbionts from the environment benefit the hosts by allowing for a degree of partner choice. According to the partner choice hypothesis, tubeworms that associate with the best-adapted partner(s) to a specific range of habitat conditions are in turn better adapted to this environment. Of course, this hypothesis assumes that there is diversity within the symbiotic partners. Phylogenetic analyses on the other hand seemed to indicate that nearly all species of vent tubeworms of the eastern Pacific were associated with the same species of symbionts: Candidatus Endoriftia persephone. However, these studies focussed on a few molecular markers. In this thesis, I used in situ hybridization and next generation sequencing to characterize the symbiont diversity at the species and strain level, as well as within individual hosts and across host species. I found that the intra-host symbiont populations are likely composed of multiple strains or lineages of the same bacterial species, that the symbiont populations separated by mid-ocean ridge discontinuities are vicariant, and that other factors such as local environmental conditions or host specificity might participate in shaping the genetic make-up of these populations. / Graduate / 0410 / 0307 / 0715 / 0306 / 0416 / 0329

Thesis (MSc)--University of Stellenbosch, 2005. / ENGLISH ABSTRACT: The Mountain Fynbos biome, a division of the Cape Floristic Region (CFR), is home to round-leafed Buchu [Agathosma betulina (Berg.) Pillans], one of South Africa’s best-known endangered herbal medicinal plants. Agathosma betulina is renowned as a traditional additive to brandy or tea, which is used for the treatment of a myriad of ailments. In its natural habitat, A. betulina thrives on mountain slopes in acid and highly leached gravelly soils, with a low base saturation and low concentrations of organic matter. To adapt to such adverse conditions, these plants have formed mutualistic symbioses with arbuscular mycorrhizal (AM) fungi. In this study, the effect of indigenous AM taxa on the physiology of A. betulina is investigated. In addition, the AM taxa responsible for these physiological responses in the plant were identified using morphological and molecular techniques. Agathosma betulina was grown under glasshouse conditions in its native rhizosphere soil containing a mixed population of AM fungi. Control plants, grown in the absence of AM fungi, were included in the experimentation. In a time-course study, relative growth rate (RGR), phosphorus (P)-uptake, P utilization cost, and carbon (C)-economy of the AM symbiosis were calculated. The data showed that the initial stages of growth were characterized by a progressive increase in AM colonization. This resulted in an enhanced P-uptake in relation to non-AM plants once the symbiosis was established. Consequently, the lower P utilization cost in AM plants indicated that these plants were more efficient in acquiring P than non-AM plants. When colonization levels peaked, AM plants had consistently higher growth respiration. This indicated that the symbiosis was resulting in a C-cost to the host plant, characterized by a lower RGR in AM plants compared to non-AM plants. Arbuscular mycorrhizal colonization decreased with increasing plant age that coincided with a decline in P-uptake and growth respiration, along with increases in RGR to a level equal to non-AM plants. Consequently, the AM benefit was only observed during the initial stages of growth. In order to identify the AM fungi in planta, morphological and molecular techniques were employed, which indicated colonization by AM fungi belonging to the genera Acaulospora and Glomus. Phylogenetic analyses of a dataset containing aligned 5.8S ribosomal RNA gene sequences from all families within the Glomeromycota, including sequences obtained during the study, supported the above mentioned identification. / AFRIKAANSE OPSOMMING: Die Fynbos bergbioom, ‘n onderafdeling van die Kaapse Floristiese Streek, huisves rondeblaar Boegoe [Agathosma betulina (Berg.) Pillans], een van Suid Afrika se bekendste bedreigde medisinale plante. Agathosma betulina is bekend vir sy gebruik as tinktuur vir die behandeling van verskeie kwale. Die plant kom voor in bergagtige streke, in suur en mineraal-arm grond, met ‘n lae organiese inhoud. Gevolglik, om aan te pas by hierdie ongunstige kondisies, vorm die plante simbiotiese assosiasies met blaasagtige, struikvormige mikorrisa (BSM). In die huidige studie is die effek van hierdie BSM op die fisiologie van A. betulina ondersoek. Die identiteit van die BSM is ook gevolglik met morfologiese en molekulêre identifikasie tegnieke bepaal. Agathosma betulina plante is onder glashuis kondisies in hul natuurlike grond gekweek, wat ‘n natuurlike populasie van BSM bevat het. Kontroles is ook in die eksperiment ingesluit en hierdie stel plante is met geen BSM geïnokuleer nie. Gevolglik is die relatiewe groeitempo, fosfor opname, fosfor verbuikerskoste asook die koolstof ekonomie van die plante bereken. Die data het getoon dat die eerste groeifase gekarakteriseer is deur toenames in BSM kolonisasie vlakke. Dit het tot ‘n hoër fosfor opname in BSM geïnokuleerde plante gelei. Die laer fosfor verbuikerskoste gedurende hierdie fase het aangedui dat die plante wat geïnokuleer is met BSM oor beter meganismes beskik het om fosfor uit die grond te bekom. Toe BSM kolonisasie vlakke gepiek het, was groei respirasie hoër in BSM geïnokuleerde plante as in die kontroles. Dit het aangedui dat die BSM kolonisasie van plante tot hoër koolstof kostes vir hierdie plante gelei het, wat weerspieël is in die laer groeitempo van die BSM geïnokuleerde plante. Die BSM kolonisasie vlakke het gedaal met toenemende ouderdom van hul gasheer plante, wat gekarakteriseer is deur ‘n laer opname van fosfor en laer groei respirasie, tesame met ‘n toename in relatiewe groeitempo tot vlakke soortgelyk aan die van die kontrole plante. Die BSM voordele vir die plant is dus net gedurende die eerste groeifase waargeneem. Die BSM wat verantwoordelik is vir hierdie fisiologiese veranderinge is gevolglik geïdentifiseer met behulp van morfologiese en molekulêre tegnieke en dit is gevind dat BSM wat behoort tot die genera Acaulospora en Glomus binne hierdie plante voorkom. Filogenetiese analise gegrond op opgelynde 5.8S ribosomale RNA geen volgordes afkomstig van al die families binne Glomeromycota asook volgordes gevind in die studie, het die bogenoemde identifikasie gestaaf.

Le facteur de transcription ZEBRA (EB1) du virus d'Epstein-Barr joue un rôle essentiel dans l'initiation de l'infection lytique et la production virale. L'ubinucléine a été identifiée comme un partenaire cellulaire de ZEBRA, capable de l'empêcher de se fixer à ses séquences d'ADN cibles. Le rôle de l'ubinucléine dans la cellule demeure inconnu, ainsi que les conséquences de son interaction avec ZEBRA dans les cellules infectées par l'EBV. Notre travail a permis, d'abord, de mieux caractériser l'ubinucléine dans la cellule épithéliale en l'identifiant comme une protéine des jonctions serrées. L'ubinucléine a été proposée comme un nouveau membre de la famille des protéines NACos (nuclear and adhesion complex components) possèdant une double localisation, les noyaux et les jonctions des cellules. Afin de mieux comprendre son rôle dans la cellule épithéliale, nous avons étudié par une approche protéomique couplée à la spectrométrie de masse les partenaires de l'ubinucléine et identifié les protéines LYRIC et RACK-1. Nos résultats suggèrent que l'ubinucléine est impliquée dans différents processus biologiques tels que la régulation de la prolifération et de l'adhésion cellulaires. Enfin, dans les cellules épithéliales infectées par l'EBV, les fonctions de l'ubinucléine semblent dépendre de sa localisation cellulaire. Au niveau nucléaire, l'ubinucléine régule négativement le cycle lytique et la production de particules virales en empêchant ZEBRA et d'autres facteurs cellulaires de se fixer à leurs promoteurs de type AP-1. Lorsqu'elle est séquestrée dans les jonctions serrées, l'inhibition de ZEBRA est levée permettant ainsi le bon déroulement du cycle lytique du virus.

Endosymbiosis is a pervasive phenomenon that has been a powerful force in insect evolution. In many well studied insect-bacterial associations, the bacteria can serve as reproductive manipulators, nutritional mutualists or defenders of their hosts. Fungi are also frequently associated with insects, and initial estimates suggest that these fungi are hyperdiverse. Saving a handful of examples, however, the functions of these fungi within insect hosts are largely unknown. This dissertation begins with a review that lays the conceptual groundwork for understanding bacterial and fungal endosymbiosis in insects. I make predictions about why one versus the other microbe might serve the insect, given any unique physiological, ecological or evolutionary conditions. I then aim to derive insights about microbial symbiosis by focusing on a particular system, that of brownbanded cockroaches, Supella longipalpa (Blattaria: Blattellidae) and their specialist wasp parasitoids, Comperia merceti (Hymenoptera: Encyrtidae). Here, I identify the symbiotic community of these two insects by using both culture-dependent and independent methods to characterize the vertically transmitted bacterial and fungal associates. Finally, I show that a heritable fungus in C. merceti, long presumed to be a mutualist, is parasitic under laboratory conditions: infected wasps incur fitness costs for housing the fungal symbiont relative to uninfected wasps. Additionally, although the fungus is not horizontally transmitted sexually, it is readily horizontally transmitted from the offspring of infected females to those of uninfected females that are using the same host.

As one of the most diverse and ecologically important clades of life, fungi are best known as pathogens, saprotrophs, mycorrhizae, and lichens. Yet an enormous amount of previously unknown diversity occurs among endophytic and endolichenic fungi--species-rich, horizontally transmitted fungi that live within asymptomatic photosynthetic structures such as leaves and lichens. Here, I explore the biodiversity of these understudied symbiotrophs and the ecological and biogeographic factors influencing their communities.To evaluate methods currently used in ecological studies of environmental samples of fungi, I assessed inter- and intraspecific divergence of a fast-evolving locus for four genera commonly found as endophytes, and compared analytical methods for identifying and delimiting OTUs. Then I used the most robust methods to show that after soil contact, seeds of a focal tree species contain diverse fungi that are closely related to endophytes and pathogens.To explore the ecological specificity of symbiotrophic fungi, I examined endophytic, endolichenic, and saprotrophic communities inhabiting physically proximate hosts in a biotically rich area of southeastern Arizona. I found that endolichenic fungi are largely distinct from plant-associated fungi, with the exception of a group of ecologically flexible symbionts that occur in lichens and mosses. Although numerous endophytes were found in non-living leaves, fungi that were highly abundant in leaf litter were seldom found as endophytes.To assess symbiotroph biodiversity and ecological specificity at a broad geographic and phylogenetic scale, I isolated>4100 endophytic and endolichenic fungi from diverse communities of plants and lichens across five climatic regions in North America. I found that the abundance, diversity, and composition of these nearly ubiquitous fungi differ as a function of climate, locality, and host. Differences among communities reflect environmental characteristics more strongly than geographic distance.Last, I addressed a series of hypotheses regarding the ecological specificity of fungi inhabiting living and non-living leaves. I show that like endophytes, saprotrophic communities are structured by environmental characteristics, and at small spatial scales by host and leaf status. Yet, differences in communities between living leaves and leaf litter suggest that most endophytes either rapidly complete their life-cycle or are out-competed by robust saprotrophs once leaves senesce.

Intimate, mutualistic, association with microbes is a common mechanism for organisms to utilize certain niches. Insects are a particularly well-studied group in this respect, frequently forming long-term, obligate associations with symbiotic microbes. These symbioses are often nutritional in nature, with the symbiont providing the host with nutrients that are otherwise unavailable. Aphids are notable for their well-defined relationship with the symbiotic Bacteria Buchnera aphidicola. By synthesizing the amino acids the aphid is unable to produce itself, Buchnera permits its host to feed on plant phloem, which lacks sufficient quantities of these essential nutrients. Buchnera, as with many obligate intracellular symbionts, has a reduced effective population size (Nₑ) due to asexual reproduction and severe population bottlenecks experienced during transmission between generations. The reduction in Nₑ has facilitated the degradation of the symbiont genome through fixation of deleterious mutations via drift. The consequences of accelerated evolutionary rates has been examined primarily through genome sequencing and comparative studies of symbionts from different host species. The work detailed in this dissertation examines the role of deleterious mutations and drift at multiple taxonomic levels. Analysis of aphid amino acid requirements utilizing an artificial diet assay revealed variation in clones of the pea aphid, Acyrthosiphon pisum. In one clone, a mutation in the arginine biosynthesis pathway appears to underlie a host dietary requirement for arginine. Examination of the number of Buchnera within an A. pisum clone also revealed variation in symbiont titer between clones. When compared across F₁ offspring of cross between a low- and a high-titer clone, extensive variation was observed in titer that exceeded variation observed in field-collected clones. No maternal effects were observed, suggesting that Buchnera is not in control of its replication. At a broader taxonomic scale, the replacement of Buchnera in the aphid Cerataphis brasiliensis was examined by sequencing the genome of its fungal symbiont (YLS). The genome of the YLS revealed a much greater metabolic capacity than Buchnera, possibly due to its extracellular habitat. The YLS exhibited signatures of elevated evolutionary rates and intron gain consistent with a reduction in Nₑ due to its symbiotic niche.

Les espèces du groupe Piptadenia sont des légumineuses endémiques du Brésil, dont la plupart sont des arbres capables de se développer sur des sols peu fertiles faisant d'eux de bons candidats pour le reboisement des terres dégradées. Les Piptadenia établissent une symbiose à la fois avec des champignons mycorhiziens à arbuscules (CMA) et des rhizobiums. Ces espèces sont proches du genre Mimosa, connu pour son affinité pour les symbiotes du genre Burkholderia. Dans ce travail de thèse nous décrivons la biodiversité des symbiotes rhizobiums associés au groupe Piptadenia, et élargissons l'affinité des Burkholderia à ce groupe de légumineuses. Les études phylogénétiques sur des marqueurs neutres et symbiotiques montrent une origine stable et ancienne de la symbiose Burkholderia/Mimoseae. Les études de spécificité d'association entre espèces de Burkholderia et espèces de Piptadenia montrent que cette dernière est lâche, les patterns d'association étant davantage liées aux sites prospectés au Brésil plutôt qu'à une sélection par l'hôte. Dans un second temps, nous avons étudié l'association tripartite entre plusieurs génotypes de Burkholderia, un CMA (Glomus clarum), et l'espèce Piptadenia gonoacantha, décrite dans la littérature comme formant une nodulation mycorhize-dépendante. Nos travaux montrent que la nodulation n'est pas CMA-dépendante, mais par contre l'efficience symbiotique des nodules dépend de la mycorhization pour certains génotypes de Burkholderia. Nous décrivons également des interactions entre symbiose rhizobienne et mycorhizienne au sein des nodules (présence du CMA dans les nodules avec sporulation dans certaines combinaisons de symbiotes). Ces travaux soulèvent la nécessité de prendre en compte les interactions génotype-génotype entre symbiotes rhizobiens et mycorhiziens lors de la sélection des inoculums dans le cadre des programmes de revégétalisation au Brésil par des arbres du groupe Piptadenia. / The Piptadenia group comprise endemic species from Brazil of which many are trees able to develop on poorly fertile soils and are good candidates for revegetation programs. Piptadenia species establish symbioses with both arbuscular mycorrhizal fungi (AMF) and rhizobia. These species are phylogenetically close to the Mimosa genus, known for its affinity for Burkholderia rhizobial symbionts. In this thesis we describe the biodiversity of rhizobial symbionts associated to the Piptadenia group, and enlarge the affinity towards Burkholderia to this group of legumes. Phylogenetic studies on neutral and symbiotic markers show a stable and ancient symbiosis Burkholderia/Mimoseae. Specificity studies between Burkholderia and Piptadenia group species show that specificity is not strong, and that patterns of associations between partners are isolation site dependent rather than linked to the host legume. In the second part of this thesis we have studied the tripartite association between several Burkholderia genotypes, an AMF (Glomus clarum), and Piptadenia gonoacantha (Pg), a legume species described as making an AMF-dependent nodulation (Jesus et al., 2005). Our experiments show that nodulation in Pg is not AMF-dependent, but that symbiotic efficiency of nodules rely on AMF presence for specific Burkholderia genotypes. We also describe interactions between rhizobial and mycorrhizal symbiosis (AMF presence in nodules, with sporulation in several symbionts combinations). Our work underlines the necessity to consider genotype-genotype interactions between rhizobial and AMF symbionts for the selection of synergistic inoculums in revegetation programs using Piptadenia group species in Brazil.

Étude moléculaire des étapes précoces de la symbiose actinorhizienne Casuarina-Frankia : analyse fonctionnelle des gènes de la plante hôte contrôlant l'infectionPlus de 80% des plantes peuvent établir une symbiose racinaire avec des champignons de l'ordre des Glomales et former des endomycorhizes à arbuscules (AM). En revanche, seules certaines espèces appartenant à dix familles d'angiospermes réunies dans le Clade des Eurosidées I peuvent établir une symbiose racinaire fixatrice d'azote. Il s'agit d'une part, des plantes de la famille des légumineuses (Fabacées) et de Parasponia associées à Rhizobium et d'autre part, des plantes actinorhiziennes associées à l'actinomycète Frankia. Comme chez les légumineuses, la symbiose actinorhizienne aboutit à la formation de nodosités (ou « nodules »), siège de la fixation d'azote par les bactéries. Cependant, contrairement aux nodules des légumineuses, le nodule actinorhizien présente une structure et un développement s'apparentant aux racines latérales. L'étude des nodosités actinorhiziennes est donc particulièrement intéressante tant pour rechercher les spécificités de cette symbiose, que pour déterminer quelles sont les caractéristiques communes avec les légumineuses. Nous avons étudié le rôle du gène CCaMK dans le processus symbiotique et l'organogenèse nodulaire chez l'arbre actinorhizien Casuarina glauca. CCaMK code pour une protéine kinase dépendante du calcium et de la calmoduline (« calcium and calmodulin dependent protein kinase »). Dans la cascade de signalisation conduisant à la nodulation et à la mycorhization chez les légumineuses, ce gène est positionné en aval des oscillations calciques (« calcium spiking ») qui ont lieu durant les premières étapes de l'interaction symbiotique. CCaMK jouerait un rôle dans la perception et le décodage des oscillations calciques, ainsi que leur transduction aux différents composants contrôlant les endosymbioses racinaires. Nous avons suivi l'expression spatio-temporelle de la fusion transcriptionnelle PromCgCCaMK::GUS au cours de la nodulation et montré que celle-ci était corrélée à la présence de Frankia tout au long du processus symbiotique, soulignant ainsi le rôle clé de CCaMK dans l'infection. Par ailleurs, nous avons cherché à déterminer l'importance du domaine autoinhibiteur de la protéine CCaMK dans l'activation du processus d'organogenèse du nodule. Pour cela, nous avons réalisé et introduit chez C. glauca des constructions géniques de CgCCaMK permettant l'expression de formes tronquées constitutivement actives, car dépourvues du domaine autoinhibiteur/CaM. Nous avons aussi utilisé des formes tronquées du gène MtCCaMK de Medicago truncatula. L'expression de ces formes tronquées de CCaMK a révélé que la levée de l'autoinhibition induit la formation de nodules spontanés indépendamment de l'actinobactérie Frankia. Les résultats obtenus suggèrent que la protéine dérégulée est capable de réactiver la voie de signalisation, ainsi que les gènes situés en aval de CCaMK, qui sont nécessaires à l'organogenèse nodulaire.Mots clés : Casuarina glauca, Frankia, CCaMK, infection, autoinhibition, nodules spontanés / Molecular study of the early stages of actinorhizal symbiosis Casuarina-Frankia: functional analysis of the host plant genes controlling the infectionMore than 80% of plant species are able to develop arbuscular mycorrhizal (AM) symbiosis in association with glomeromycete fungi. In contrast, only some species of the Eurosid I clade, confined to four orders and ten Angiosperm families, are able to form nitrogen-fixing root nodule symbioses with soil bacteria. This concerns plants of the legume family (Fabaceae) and Parasponia associated with Rhizobium bacteria and actinorhizal plants associated with the actinomycete Frankia. Similarly to Legumes, the actinorhizal symbiosis results in the formation of nitrogen-fixing root nodules. However, unlike legume nodule, the actinorhizal nodule has a same origin and structure than a lateral root. Thus, the study of actinorhizal nodules is of particular interest not only for investigating its specific properties but also, for determining common characteristics shared with legume nodules.We have studied the role of CgCCaMK gene during the symbiotic process and nodule organogenesis in the actinorhizal tree Casuarina glauca. CCaMK encodes a calcium and calmodulin dependent protein kinase. In the signalisation cascade leading to both nodulation and mycorrhization in legumes, this gene is acting downstream the calcium oscillations (« calcium spiking ») that occur during the early steps of the symbiotic interaction. It has been suggested that these calcium oscillations are decoded and transduced by the CCaMK protein.We have monitored the spatio-temporal expression of a PromCgCCaMK::GUS fusion during actinorhizal nodulation and have shown that reporter gene expression was correlated with the presence of Frankia along the symbiotic process. This data highlights the role of CgCCaMK during Frankia infection. In addition, we have investigated the role of the CCaMK autoinhibitory/CaM domain in actinorhizal nodule organogenesis. To achieve this goal, we have obtained truncated versions of CgCCaMK lacking the autoinhibitory/CaM domain, and then expressed them into C. glauca. We have also used truncated forms of MtCCaMK from Medicago truncatula. The expression of these CCaMK constructs from C. glauca and M. truncatula was found to induce spontaneous nodulation in the absence of Frankia bacteria. These results suggest that deregulation of the calcium and calmodulin dependent protein kinase is able to reactivate the symbiotic signalling pathway and genes acting downstream CCaMK that are needed for nodule organogenesis.Key words: Casuarina glauca, Frankia, CCaMK, infection, autoinhibition, spontaneous nodules

This diploma thesis has focused on the diversity and biogeography of Asterochloris photobionts. Since no study so far has been published on the biogeography of symbiotic microorganisms, the presented thesis is the first attemp to trace the biogeographic distribution and endemism of symbionts. By gathering 121 Asterochloris sequences obtained from lichen thalli sampled outside Europe and America, the diversity within the genus increased dramatically. The phylogenetic analysis based on the concatenated alignment of ITS rDNA and actin sequences obtained from Cladonia and Stereocaulon photobionts revealed 28 differently supported clades. Of them, eight lineages were newly discovered. Three environmental factors explaining the best the distribution pattern of Asterochloris photobionts were selected according to the statistical tests of the phylogenetic signal: two different types of biogeographical ecoregions and the substrate type. In general, the genus Asterochloris is distributed cosmopolitally, with a very low rate of endemism. Newly obtained data indicate that the restricted distribution of any photobiont clade is not caused by either historic or biological factors, but more likely by specific climatic or habitat preferences.