The relative impact of lichen symbiotic partners to repeated copper uptake

The relative impact of lichen photobiont and mycobiont was evaluated by submitting nine lichen species with: (i) different photobiont types; (ii) different lichen growth forms; and (iii) different nutrients, pH, humidity preferences; to a range of Cu concentrations (μM) supplied in repeated cycles to simulate the natural process of uptake under field conditions. The physiological performance of the photosystem II photochemical reactions was measured using F_v/F_m and the metabolic activity of the mycobiont was evaluated using ergosterol and intracellular K-loss as indicators. Lichens with higher cation exchange capacity showed higher intracellular Cu uptake and their ecology seemed to be associated with low-nutrient environments. Thus the wall and external matrix, mainly characteristic of the mycobiont partner, cannot be ignored as the first site of interaction of metals with lichens. No common intracellular Cu concentration threshold was found for the physiological impacts observed in the different species. Most physiological effects of Cu uptake in sensitive lichens occurred for intracellular Cu below 200 μg/g dw whereas more tolerant species were able to cope with intracellular Cu at least 3 times higher. Cyanobacterial lichens showed to be more sensitive to Cu uptake than green-algal lichens. Within the Trebouxia lichens, different species showed different sensitivities to Cu uptake, suggesting that the mycobiont may change the microenvironment close to the photobiont partner providing different degrees of protection. Despite the fact that the photobiont is the productive partner, the metabolic activity of the mycobiont of lichen species adapted to environments rich in nutrients, showed to be more sensitive to Cu uptake than the photochemical performance of the photobiont.     

Alterations in secondary metabolism of aposymbiotically grown mycobionts of Xanthoria elegans and cultured resynthesis stages

HPLC analyses of Xanthoria elegans cultivated on different media and either aposymbiontically or with its photobiont revealed that the carbon source and the presence of the algal partner have an impact on the secondary metabolism of the mycobiont. The aposymbiotically (without photobiont) grown mycobiont contained up to 70% more of the main compounds in its thallus than in resynthesis stage. Although this is speculative, the induction of the polyketide pathway may be a feedback mechanism to the absence of the photobiont. All cultures produce a variety of substances which were not detectable in the voucher specimen. Besides physcion (the major substance), we were able to identify emodin as well as physcion-bisanthrone, teloschistin monoacetate and derivatives. A strong inducible effect on the production of physcion, physcion-bisanthrone and on their precursors and derivatives was found for mannitol. By contrast, supplementation of ribitol had negligible effects, if any, on polyketide quantities although it is the main carbon source for the mycobiont in free-living lichens with Trebouxia photobiont.     

Zn/Pb-tolerant lichens with higher content of secondary metabolites produce less phytochelatins than specimens living in unpolluted habitats

Many lichens can cope with heavy-metal stress, however, the mechanisms of lichen tolerance are still not fully understood. Some lichen secondary metabolites (depsides and depsidones), produced in lichens by the fungal symbiont and accumulated on the outer surface of its hyphae, are supposed to play an important role in the extracellular immoblilization of heavy metals. Lichen photobionts (algal partners in the symbiosis), although surrounded by the mycobiont hyphae, may also accumulate high amounts of trace metals. This can lead to physiological disruptions and morphological damage in algal cells and hence affect the lichen physiological status. We hypothesized that lichen species/specimens living in heavily polluted sites and showing HM tolerance possess a higher content of secondary metabolites than those living in unpolluted sites. Hence, their photobionts can be better protected from the excess of metal ions and need to produce less metal-complexing phytochelatins (PCn) to combat metal toxicity. Specimens of Hypocenomyce scalaris, Cladonia furcata and Lepraria spp. sampled from Zn/Pb-polluted and control sites were compared for the accumulation of Zn/Pb and secondary metabolites, as well as for their production of phytochelatins and glutathione in response to experimental Zn or Pb exposure. Generally, the lichen specimens sampled from the HM-polluted site contained higher amounts of Zn and Pb as well as lichen substances (different depsides and depsidones) than those from the control site. A strong positive correlation was found between the accumulation of secondary metabolites and Zn/Pb accumulation (R² = 0.98 and 0.63, respectively). For the first time, production of phytochelatins (PC_2-3) in response to Zn and Pb (50-200 μM) exposure was found in H. scalaris, L. elobata, L. incana and C. furcata. In both species of Lepraria also cysteine, a substrate for GSH and PCs synthesis was detected. The lichens from the polluted site produced under the same exposure conditions, or in response to higher metal concentrations, lower amounts of PCn than those sampled from the control site. It strongly suggests that less Zn and Pb ions reached the photobiont cells of the lichens containing higher amounts of secondary metabolites (lecanoric, fumarprotocetraric, stictic, constictic acids, antranorin). The results obtained support the putative role of some metabolites in heavy-metal tolerance of the lichens inhabiting metal-polluted habitats.     

Response to copper stress in aposymbiotically grown lichen mycobiont Cladonia cristatella: uptake,viability, ergosterol and production of non-protein thiols

The mycobiont of lichens usually determines the morphology of the symbiotic organism and is also dominates in terms of biomass. However, its role for sensitivity or tolerance of lichens to heavy metals is almost unknown. In the present study, the influence of copper (Cu) on the aposymbiotically-grown mycobiont of Cladonia cristatella was assessed. Intracellular Cu uptake was correlated with increasing Cu concentrations over a 24-h exposure time. Viability, measured as the degree of reduction of triphenyltetrazolium chloride to triphenyl formazan, as well as to ergosterol levels, decreased with growing Cu concentrations tested. Reduced glutathione (GSH) was found to be the most abundant low-molecular-weight thiol in the hyphae of C. cristatella and its intracellular content increased at concentrations of 10 μm Cu. Higher Cu concentrations caused a significant decrease in GSH, possibly due to heavy metal-induced oxidation of GSH to glutathione disulphide (GSSG). Free cysteine levels were relatively constant. As expected, we did not observe the production of phytochelatins in the mycobiont, contrary to what is found in intact lichens and axenic cultures of their photobionts.     

The impact of nitrogen deposition on photobiont‐mycobiont balance of epiphytic lichens in subtropical forests of central China

Excessive nitrogen (N) deposition can impact lichen diversity in forest ecosystems, and this is a particular situation in China. Here, we examined the N uptake, assimilation, and the impact of excessive N deposition on the symbiotic balance of dominant epiphytic lichens in the subtropical forests in the Mts. Shennongjia of central China. The results show that lichen species took up, assimilated and utilized more ammonium than nitrate in a species‐specific way, following the increase of N availability. The photobiont of the lichens decreased with the increase of N concentration following an initial increase, while the mycobiont response to the N addition was not apparent. Considerable variation in response to excessive N deposition exists among the lichen species. Usnea longissima could regulate its N uptake, resulting in a stable photobiont‐mycobiont ratio among N treatments. In contrast, the photobiont‐mycobiont ratio of other four lichens increased initially but decreased when N concentration exceeded a certain level, and N stress may have broken the balance between photobiont and mycobiont of these lichens. Our results suggest that most epiphytic lichens in subtropical forest of central China could uptake and assimilate more ammonium than nitrate and that the balance between photobiont and mycobiont of many epiphytic lichens might change with the increasing N deposition load, which could impact the lichen diversity of this forest ecosystem.     

Cyanolichens can have both cyanobacteria and green algae in a common layer as major contributors to photosynthesis

Background and Aims

Cyanolichens are usually stated to be bipartite (mycobiont plus cyanobacterial photobiont). Analyses revealed green algal carbohydrates in supposedly cyanobacterial lichens (in the genera Pseudocyphellaria, Sticta and Peltigera). Investigations were carried out to determine if both cyanobacteria and green algae were present in these lichens and, if so, what were their roles.

Methods

The types of photobiont present were determined by light and fluorescence microscopy. Small carbohydrates were analysed to detect the presence of green algal metabolites. Thalli were treated with selected strengths of Zn²⁺ solutions that stop cyanobacterial but not green algal photosynthesis. CO₂ exchange was measured before and after treatment to determine the contribution of each photobiont to total thallus photosynthesis. Heterocyst frequencies were determined to clarify whether the cyanobacteria were modified for increased nitrogen fixation (high heterocyst frequencies) or were normal, vegetative cells.

Key Results

Several cyanobacterial lichens had green algae present in the photosynthetic layer of the thallus. The presence of the green algal transfer carbohydrate (ribitol) and the incomplete inhibition of thallus photosynthesis upon treatment with Zn²⁺ solutions showed that both photobionts contributed to the photosynthesis of the lichen thallus. Low heterocyst frequencies showed that, despite the presence of adjacent green algae, the cyanobacteria were not altered to increase nitrogen fixation.

Conclusions

These cyanobacterial lichens are a tripartite lichen symbiont combination in which the mycobiont has two primarily photosynthetic photobionts, ‘co-primary photobionts’, a cyanobacterium (dominant) and a green alga. This demonstrates high flexibility in photobiont choice by the mycobiont in the Peltigerales. Overall thallus appearance does not change whether one or two photobionts are present in the cyanobacterial thallus. This suggests that, if there is a photobiont effect on thallus structure, it is not specific to one or the other photobiont.     

Hardening by partial dehydration and ABA increase desiccation tolerance in the cyanobacterial lichen Peltigera polydactylon

* BACKGROUND AND AIMS: The ability of partial dehydration and abscisic acid pretreatments to increase desiccation tolerance in the cyanobacterial lichen Peltigera polydactylon was tested. * METHODS: Net photosynthesis and respiration were measured using infrared gas analysis during a drying and rehydration cycle. At the same time, the efficiency of photosystem two was measured using chlorophyll fluorescence, and the concentrations of chlorophyll a were spectrophotometrically assayed. Heat production was also measured during a shorter drying and rehydration cycle using differential dark microcalorimetry. * KEY RESULTS: Pretreating lichens by dehydrating them to a relative water content of approx. 0.65 for 3 d, followed by storing thalli hydrated for 1 d in the light, significantly improved their ability to recover net photosynthesis during rehydration after desiccation for 15 but not 30 d. Abscisic acid pretreatment could substitute for partial dehydration. The improved rates of photosynthesis during the rehydration of pretreated material were not accompanied by preservation of photosystem two activity or chlorophyll a concentrations compared with untreated lichens. Partial dehydration and ABA pretreatments appeared to have little direct effect on the desiccation tolerance of the mycobiont, because the bursts of respiration and heat production that occurred during rehydration were similar in control and pretreated lichens. * CONCLUSIONS: Results indicate that the photobiont of P. polydactylon possesses inducible tolerance mechanisms that reduce desiccation-induced damage to carbon fixation, and will therefore improve the supply of carbohydrates to the whole thallus following stress. In this lichen, ABA is involved in signal transduction pathways that increase tolerance of the photobiont.     

Assessing intraspecific diversity in a lichen-forming fungus and its green algal symbiont: Evaluation of eight molecular markers

To facilitate marker selection in sequence-based studies on genetic diversity and symbiont selectivity in lichens we conducted a comparison of eight molecular markers in the lichen-forming fungus Lasallia pustulata and its trebouxioid photobiont. We compared mtSSU rDNA, mtLSU rDNA, MCM7, TSR1 (mycobiont) and nrITS rDNA, COX2, psbJ-L intergenic spacer, rbcL (photobiont) of 45 individuals from European populations of L. pustulata. Mycobiont and photobiont loci had congruent phylogenetic signals. Based on the results of this study we recommend the use of MCM7 and TSR1 (mycobiont), and nrITS rDNA and COX2 (photobiont). In this specific study system we found no sequence variability in the mycobiont loci EF1, nrITS rDNA, RPB1, and RPB2, which we sequenced for a subset of individuals. We had limited success amplifying GPD (mycobiont), actin and chloroplast LSU rDNA (photobiont), however, we do not rule out that these loci could be valuable markers in other species.     

Lead and copper effects on lipid metabolism in cultured lichen photobionts with different phosphorus status

We evaluated the ability of heavy metals (copper, lead) to alter lipid metabolism in four algal lichen photobionts following short term exposure. Metal concentrations (10 microM) were equivalent to environmentally relevant levels that have been reported to have effects on intact algae. The algae were grown under normal or deficient phosphate conditions to assess any interactions with the heavy metal stress. Given the frequent sensitivity of lichens to copper and lead, there were surprisingly small changes on lipid metabolism, as assessed by radiolabelling from [1-14C]acetate. The main effects, which were seen in a number of cases, were an overall inhibition of total lipid labelling and a relative increase in the labelling of triacylglycerols in the non-polar fraction. Both of these changes can be viewed as reflecting general toxicity of heavy metals. The Coccomyxa photobiont species were more sensitive than Trebouxia species, which fits with the general distribution of the latter in lichens inhabiting harsh environments.     

Symbiosis as a successful strategy in continental Antarctica: performance and protection of Trebouxia photosystem II in relation to lichen pigmentation

Lichens as symbiotic associations consisting of a fungus (the mycobiont) and a photosynthetic partner (the photobiont) dominate the terrestrial vegetation of continental Antarctica. The photobiont provides carbon nutrition for the fungus. Therefore, performance and protection of photosystem II is a key factor of lichen survival. Potentials and limitations of photobiont physiology require intense investigation to extend the knowledge on adaptation mechanisms in the lichen symbiosis and to clarify to which extent photobionts benefit from symbiosis. Isolated photobionts and entire lichen thalli have been examined. The contribution of the photobiont concerning adaptation mechanisms to the light regime and temperature conditions was examined by chlorophyll a fluorescence and pigment analysis focusing on the foliose lichen Umbilicaria decussata from North Victoria Land, continental Antarctica. No photoinhibition has been observed in the entire lichen thallus. In the isolated photobionts, photoinhibition was clearly temperature dependent. For the first time, melanin in U. decussata thalli has been proved. Though the isolated photobiont is capable of excess light protection, the results clearly show that photoprotection is significantly increased in the symbiotic state. The closely related photobiont of Pleopsidium chlorophanum, a lichen lacking melanin, showed a higher potential of carotenoid-based excess light tolerance. This fact discriminates the two photobionts of the same Trebouxia clade. Based on the results, it can be concluded that the successful adaptation of lichens to continental Antarctic conditions is in part based on the physiological potential of the photobionts. The findings provide information on the success of symbiotic life in extreme environments.     

Parietin in the tolerant lichen Xanthoria parietina (L.) Th. Fr. increases protection of Trebouxia photobionts from cadmium excess

Secondary metabolites of lichens can be involved in production of chelates with heavy metals. We hypothesized that parietin plays important role in protection of photobiont cells in Xanthoria parietina from an excess of cadmium ions. Two types of X. parietina lichen thalli, natural with presence of secondary metabolite parietin (p+) as well as without parietin (p−) were exposed to different doses of cadmium (up to 300 μmol g⁻¹ dw). Based on determination of the total and intracellular Cd-accumulation, ergosterol and thiobarbituric acid reactive substances (TBARS) content did not show statistically significant differences in the response of both types of thalli (p+ and p−). However, a stronger toxic effect of the highest Cd-dose on photosynthetic pigment content and chlorophyll a fluorescence was observed in the parietin-depleted thalli. The protective role of parietin against Cd excess was better supported and concluded from the differences observed in the production of non-protein thiol compounds (cysteine, glutathione and phytochelatins) involved in Cd detoxification. In the p+ thalli Cys content was stable but GSH content slightly decreased in the studied Cd range, while in the p− thalli these compounds were completely absent at high Cd doses. At Cd doses higher than 37.5 μmol Cd g⁻¹ dw, toxic to both types of X. parietina thalli, Cys and GSH contents were significantly higher in p+ than in p− thalli. Also, the photobiont partner in the p+ thalli was better protected of the metal exposition, and able to produce phytochelatins (PCs) over the whole range of metal, while in the p− thalli the production was completely inhibited at 75 μmol Cd g⁻¹ dw and higher concentrations, together with the inhibition of cysteine (Cys) and reduced glutathione (GSH) production. The obtained results indicate that the parietin layer is a natural barrier decreasing Cd access to algal cells in X. parietina. Comparison of PCs production appeared to be the most sensitive marker for estimation of Cd availability to photobiont in the symbiotic system.     

Carbon and nitrogen distribution in the green algal lichens<Emphasis Type="Italic"> Hypogymnia physodes</Emphasis> and<Emphasis Type="Italic"> Platismatia glauca</Emphasis> in relation to nutrient supply

With the aim of understanding how some lichens can survive intensive fertilization we investigated two green algal ( Trebouxia) lichens, Hypogymnia physodes (L.) Nyl. and Platismatia glauca (L.) W. Culb., and compared control (Ctr), and intensively fertilized (F) thalli. We measured total N, proteins and amino acids to assess lichen N status. Chlorophyll a indicated photosynthetic capacity and photobiont mass, ergosterol the metabolic demands of the fungus, and chitin the fungal biomass. For carbon status we measured glucose, the photobiont ( Trebouxia) export product ribitol, and the mycobiont-specific carbohydrates arabitol and mannitol. The F-thalli had 2-3 times higher protein and N concentrations, 5-10 times higher chlorophyll a concentrations, while ergosterol and chitin were doubled. The ribitol concentrations were 4-5 times higher in the F-thalli, while the fungal carbohydrates did not increase to the same extent. The amino acid arginine had increased 60-fold. The F-thalli also had a relatively higher N investment in the photobiont in relation to mycobiont tissue compared to the Ctr-thalli, probably resulting in an increased capacity for carbon assimilation, most possibly required for maintaining the higher nutrient status of the F-thalli. Arginine accumulation possibly avoided toxic effects of accumulated NH4+, albeit binding a significant fraction of assimilated carbon.     

Population structure of mycobionts and photobionts of the widespread lichen Cetraria aculeata

Lichens are symbioses between fungi (mycobionts) and photoautotrophic green algae or cyanobacteria (photobionts). Many lichens occupy large distributional ranges covering several climatic zones. So far, little is known about the large‐scale phylogeography of lichen photobionts and their role in shaping the distributional ranges of lichens. We studied south polar, temperate and north polar populations of the widely distributed fruticose lichen Cetraria aculeata. Based on the DNA sequences from three loci for each symbiont, we compared the genetic structure of mycobionts and photobionts. Phylogenetic reconstructions and Bayesian clustering methods divided the mycobiont and photobiont data sets into three groups. An amova shows that the genetic variance of the photobiont is best explained by differentiation between temperate and polar regions and that of the mycobiont by an interaction of climatic and geographical factors. By partialling out the relative contribution of climate, geography and codispersal, we found that the most relevant factors shaping the genetic structure of the photobiont are climate and a history of codispersal. Mycobionts in the temperate region are consistently associated with a specific photobiont lineage. We therefore conclude that a photobiont switch in the past enabled C. aculeata to colonize temperate as well as polar habitats. Rare photobiont switches may increase the geographical range and ecological niche of lichen mycobionts by associating them with locally adapted photobionts in climatically different regions and, together with isolation by distance, may lead to genetic isolation between populations and thus drive the evolution of lichens.     

Photobiont selectivity for lichens and evidence for a possible glacial refugium in the Ross Sea Region, Antarctica

Lichens are a symbiosis consisting of heterotrophic, fungal (mycobiont) and photosynthetic algal or cyanobacterial (photobiont) components. We examined photobiont sequences from lichens in the Ross Sea Region of Antarctica using the internal transcribed spacer region of ribosomal DNA and tested the hypothesis that lichens from this extreme environment would demonstrate low selectivity in their choice of photobionts. Sequence data from three targeted lichen species (Buellia frigida, Umbilicaria aprina and Umbilicaria decussata) showed that all three were associated with a common algal haplotype (an unnamed Trebouxia species) which was present in all taxa and at all sites, suggesting lower selectivity. However, there was also association with unique, local photobionts as well as evidence for species-specific selection. For example, the cosmopolitan U. decussata was associated with two photobiont species, Trebouxia jamesii and an unnamed species. The most commonly collected lichen (B. frigida) had its highest photobiont haplotype diversity in the Dry Valley region, which may have served as a refugium during glacial periods. We conclude that even in these extreme environments, photobiont selectivity still has an influence on the successful colonisation of lichens. However, the level of selectivity is variable among species and may be related to the ability of some (e.g. B. frigida) to colonise a wider range of habitats.     

Fungal farmers or algal escorts: lichen adaptation from the algal perspective

Domestication of algae by lichen‐forming fungi describes the symbiotic relationship between the photosynthetic (green alga or cyanobacterium; photobiont) and fungal (mycobiont) partnership in lichen associations ( Goward 1992 ). The algal domestication implies that the mycobiont cultivates the alga as a monoculture within its thallus, analogous to a farmer cultivating a food crop. However, the initial photobiont ‘selection’ by the mycobiont may be predetermined by the habitat rather than by the farmer. When the mycobiont selects a photobiont from the available photobionts within a habitat, the mycobiont may influence photobiont growth and reproduction ( Ahmadjian & Jacobs 1981 ) only after the interaction has been initiated. The theory of ecological guilds ( Rikkinen et al. 2002 ) proposes that habitat limits the variety of photobionts available to the fungal partner. While some studies provide evidence to support the theory of ecological guilds in cyanobacterial lichens ( Rikkinen et al. 2002 ), other studies propose models to explain variation in symbiont combinations in green algal lichens ( Ohmura et al. 2006 ; Piercey‐Normore 2006 ; Yahr et al. 2006 ) hypothesizing the existence of such guilds. In this issue of Molecular Ecology, Peksa & ?kaloud (2011) test the theory of ecological guilds and suggest a relationship between algal habitat requirements and lichen adaptation in green algal lichens of the genus Lepraria. The environmental parameters examined in this study, exposure to rainfall, altitude and substratum type, are integral to lichen biology. Lichens have a poikilohydric nature, relying on the availability of atmospheric moisture for metabolic processes. Having no known active mechanism to preserve metabolic thallus moisture in times of drought, one would expect a strong influence of the environment on symbiont adaptation to specific habitats. Adaptation to changes in substrata and its properties would be expected with the intimate contact between crustose lichens in the genus Lepraria. Altitude has been suggested to influence species distributions in a wide range of taxonomic groups. This is one of the first studies to illustrate an ecological guild, mainly for exposure to rainfall (ombrophiles and ombrophobes), with green algal lichens.     

Asymmetric Co-evolution in the Lichen Symbiosis Caused by a Limited Capacity for Adaptation in the Photobiont

It is proposed that lichen photobionts, compared to mycobionts, have very limited capacity to evolve adaptations to lichenization, so that the symbionts in lichens do not co-evolve. This is because lichens have (a) no sequential selection of photobiont cells from one lichen into another needed for Darwinian natural selection and (b) no photobiont sexual reproduction in the thallus. Molecular studies of lichen photobionts indicate no predictable patterns of photobiont lineages that occur in lichens so supporting this proposal. Any adaptation by photobionts accumulating beneficial mutations for lichenization is probably insignificant compared to the rate of mycobiont adaptation. This proposal poses questions for research relating the photobiont sexual cycle (genetic and cellular), the fate of photobiont lineages after lichenization, whether lineages of photobionts in thalli change with time, thallus formation by from spores as well as carbohydrate movement from photobionts to mycobionts and regulation of co-development of the symbionts in the thallus.     

Influences of Nitrogen Sources on Usnic Acid Production in a Cultured Mycobiont of the Lichen Usnea hirta (L.) Wigg.

Effects of the nitrogen sources in the medium for the production of secondary metabolites in lichens were examined. The usnic acid production by a mycobiont of the lichen Usnea hirta was higher in the liquid medium containing ammonium and nitrate ions than in those containing amino acids.     

Influences of nitrogen sources on usnic acid production in a cultured Mycobiont of the lichen Usnea hirta (L.) Wigg

Effects of the nitrogen sources in the medium for the production of secondary metabolites in lichens were examined. The usnic acid production by a mycobiont of the lichen Usnea hirta was higher in the liquid medium containing ammonium and nitrate ions than in those containing amino acids.     

Lichen responses to nitrogen and phosphorus additions can be explained by the different symbiont responses

? Responses to simulated nitrogen (N) deposition with or without added phosphorus (P) were investigated for three contrasting lichen species - the N-sensitive Alectoria sarmentosa, the more N-tolerant Platismatia glauca and the N(2) -fixing Lobaria pulmonaria- in a field experiment. ? To examine whether nutrient limitation differed between the photobiont and the mycobiont within the lichen, the biomass responses of the respective bionts were estimated. ? The lichenized algal cells were generally N-limited, because N-stimulated algal growth in all three species. The mycobiont was P-limited in one species (A. sarmentosa), but the growth response of the mycobionts was complex, as fungal growth is also dependent on a reliable carbon export from the photobiont, which may have been the reason for the decrease of the mycobiont with N addition in P. glauca. ? Our findings showed that P availability was an important factor when studying effects of N deposition, as P supply can both mitigate and intensify the negative effects of N on epiphytic lichens.     

Recognition mechanisms during the pre-contact state of lichens: I. Mycobiont-photobiont interactions of the mycobiont of Fulgensia bracteata

Lichens are an association of a photoautotrophic alga/cyanobacteria (photobiont) and a heterotrophic fungus (mycobiont) constituting the lichen thallus as a complex phenotype. Many mycobionts reproduce sexually and the ascospores are dispersed without the photobiont. For successful re-lichenization the specific photobiont must be recognized, contacted, and incorporated by the mycobiont. A so-called pre-contact stage has been postulated as the initial step of a gradual recognition process. In the present study, the effect of the specific Trebouxia photobiont, an unspecific Asterochloris photobiont and the non-lichenizing green alga Myrmecia bisecta on the development of the mycobiont Fulgensia bracteata was assessed by pre-contact assays. Three hypotheses were confirmed: (i) the pre-contact stage exists, (ii) it is characterized by morphological reactions in the development of the mycobiont, and (iii) the reactions depend on the interacting alga. Control conditions revealed a mycelial growth arrest but this effect was not observed in the presence of any of the three algae. Different algae induce distinct growth patterns with respect to hyphal length, morphological characteristics, and formation of mucilage. The specific Trebouxia photobiont had a positive impact on hyphal growth, branching frequency, and mucilage formation. These effects were less explicit with the non-specific Asterochloris photobiont. Myrmecia bisecta induced uncharacteristic growth patterns with pronounced hyphal growth and high numbers of aerial hyphae but less formation of mucilage. These results indicate that symbiont recognition mechanisms are established before physical contact. Pre-contact reactions may be an evolutionary advantage that supports the persistence of the mycobiont on newly colonized sites and improves the probability of re-lichenization.