The phenolic compounds in <Emphasis Type="Italic">Cladonia</Emphasis> lichens are not antimicrobial in soils

According to classic text books on lichen biology, the phenolic secondary chemicals in lichens have antibiotic effects on soil microorganisms and mycorrhizal fungi in ecosystems. However, the experimental evidence for this under natural conditions is still relatively scarce. We examined some of the assumptions behind the concept of antimicrobial effects of lichen secondary substances: (1) the secondary substances of Cladonia stellaris, usnic and perlatolic acids, are leached out from the lichens by rainwater; (2) these substances inhibit the microbial activity of soil, and; (3) since they are extremely resistant to microbial decomposition, the soil underneath a continuous lichen mat is enriched in usnic and perlatolic acids. Our results did not support any of these assumptions. The evidence for the antimicrobial activity of lichen secondary substances seems to be weak in comparison to other suggested functions such as light filtering and herbivore protection. We suggest that it is time to re-evaluate the evidence for the antimicrobial ecological role of lichen secondary substances in natural systems.     

Dissociation and metal-binding characteristics of yellow lichen substances suggest a relationship with site preferences of lichens

Background and Aims: Many species of lichen-forming fungi contain yellow or orangeextracellular pigments belonging to the dibenzofurans (usnicacid), anthraquinones (e.g. parietin) or pulvinic acid group.These pigments are all equally efficient light screens, leadingus to question the potential ecological and evolutionary significanceof diversity in yellow and orange lichen substances. Here thehypothesis is tested that the different pigments differ in metal-bindingcharacteristics, which suggest that they may contribute to adaptationto sites differing in pH and metal availability. Methods: UV spectroscopy was used to study the dissociation and the pHdependence of the metal-binding behaviour of seven isolatedlichen substances in methanol. Metals applied were selectedmacro- and micro-nutrients (Cu²⁺, Fe²⁺, Fe³⁺, Mg²⁺, Mn²⁺ andZn²⁺). Key Results: All the pigments studied are strong to moderate acids with pK_a1values between 2·8 and 4·5. Metal complexationis common in the lichen substances studied. Complexation takesplace under acidic conditions with usnic acid, but under alkalineconditions with parietin and most compounds of the pulvinicacid group. The pulvinic acid derivative rhizocarpic acid formsmetal complexes both in the acidic and the alkaline range. Conclusions: Metal complexation by lichen substances could be a prerequisitefor lichen substance-mediated control of metal uptake. Assumingsuch an effect at pH values where the affinity of the metalfor the lichen substance is intermediate would explain the strongpreference of lichens with usnic or rhizocarpic acids to acidicsubstrata. Moreover, it would explain the preference of lichenswith parietin and some lichens with compounds of the pulvinicacid group either for nutrient-rich substrata at low pH or forcalcareous substrata.     

Secondary chemistry of cultured mycobionts: formation of a complete chemosyndrome by the lichen fungus of Lobaria spathulata

Abstract:The study aimed to optimize culture conditions and nutrient requirements for the production of secondary metabolites by the cultured mycobiont Lobaria spathulata. This species proved to be an excellent model system for such studies, as the complete chemosyndrome found in the natural lichen thallus was repeatedly formed in the cultured mycobiont with differentiated, aerial mycelia. Nutrient media containing the disaccharide, sucrose, were found to favour both rapid growth and the production of typical lichen substances. Higher proportions of the secondary compounds were detected in the developing mycobiont than in mature lichen thalli.     

Lichen substance concentrations in the lichen Hypogymnia physodes are correlated with heavy metal concentrations in the substratum

Lichen substances (i.e. lichen-specific carbon-based secondary compounds) are known to be involved in the uptake and immobilization of metal ions, though the biochemical mechanisms of this interaction are largely unexplained. Previous research on potential effects of lichen substances on heavy metal uptake and tolerance mostly focused on lichens in heavily polluted areas with exceptionally high metal concentrations. In the present study, we aimed at gathering information as to whether lichen substances might be involved in the fine-tuning of metal uptake even at not or low-polluted sites. Therefore, we studied lichen substance concentrations in the epiphytic lichen Hypogymnia physodes and metal concentrations in its substratum in a montane spruce forest of Germany. H. physodes produces two depsides and five depsidones, which had been shown to be involved in metal homeostasis, namely in Cu and Mn uptake, in previous laboratory experiments. The amount of lichen substances increased with increasing heavy metal concentration in the substratum, though the latter varied only in the range of a few μmol g⁻¹ between the sample trees. Variability of lichen substance concentrations in H. physodes within the individual trees was low. Among the different lichen substances of H. physodes, the amount of the depsidone physodalic acid relative to the total of lichen substances was most closely correlated to the concentrations of Cu and Mn in the substratum, whereas the amount of the depsidone 3-hydroxyphysodic acid decreased both with increasing concentrations of these two metals and physodalic acid. Thus, our data suggest that lichen substances contribute to metal homeostasis not only in heavy metal-rich habitats, but also at not or low-polluted sites where the lichen substances apparently help to maintain constant intracellular metal concentrations despite of spatially varying availabilities of metal ions.     

The secondary metabolism of Arabidopsis thaliana: growing like a weed

Despite its small stature, short life-cycle and highly reduced genome, Arabidopsis thaliana has a complement of secondary metabolites that is every bit as numerous and diverse as those of other plant taxa. The list of secondary metabolites isolated from this model species has expanded more than five-fold in the past ten years, and many more substances are likely to be added in the near future. Among the classes of compounds recently discovered are coumarins, benzenoids and terpenoids. Many A. thaliana secondary metabolites appear to have internal roles within the plant instead of (or in addition to) mediating ecological interactions.     

Does secondary chemistry enable lichens to grow on iron-rich substrates?

Lichen substances are shown to increase or to inhibit the adsorption of Fe at cation exchange sites. The influence on the adsorption strongly differs between individual lichen substances and is different for Fe²⁺ and Fe³⁺. These results add a new biological role to the known functions of lichen secondary metabolites. In an experiment with cellulose filters, which were soaked with acetone solutions of lichen substances and were then incubated with micromolar solutions of FeCl₂ or FeCl₃, many lichen substances were found to increase Fe³⁺ adsorption, whereas others had no effect. Most lichen substances had no effect on Fe²⁺ adsorption, but two were found to reduce and one to increase the level of adsorption. Lichens of Fe-poor and -rich sites contain lichen substances with different adsorption behavior towards Fe²⁺ and Fe³⁺. All the studied lichen substances, which only occur in lichens of Fe-poor sites, turned out to be effective Fe³⁺ adsorbents. Lichens of Fe-bearing rock and slag, however, were found to lack lichen substances, or to contain substances that did not adsorb Fe³⁺ and had no effect on Fe²⁺ adsorption, or thirdly, to contain substances that increased Fe³⁺ adsorption, but decreased Fe²⁺ adsorption. These results suggest that lichen substances do play a significant role in Fe adsorption in lichens and determine their tolerance to excess concentrations of Fe. Notwithstanding the strong correlation between the secondary chemistry of lichen species and their preference for Fe-rich or Fe-poor substrates, the postulated mechanism of temporary Fe adsorption by lichen substances has to be subject of future biochemical research.     

Extremotolerance and Resistance of Lichens: Comparative Studies on Five Species Used in Astrobiological Research II. Secondary Lichen Compounds

Lichens, which are symbioses of a fungus and one or two photoautotrophs, frequently tolerate extreme environmental conditions. This makes them valuable model systems in astrobiological research to fathom the limits and limitations of eukaryotic symbioses. Various studies demonstrated the high resistance of selected extremotolerant lichens towards extreme, non-terrestrial abiotic factors including space exposure, hypervelocity impact simulations as well as space and Martian parameter simulations. This study focusses on the diverse set of secondary lichen compounds (SLCs) that act as photo- and UVR-protective substances. Five lichen species used in present-day astrobiological research were compared: Buellia frigida, Circinaria gyrosa, Rhizocarpon geographicum, Xanthoria elegans, and Pleopsidium chlorophanum. Detailed investigation of secondary substances including photosynthetic pigments was performed for whole lichen thalli but also for axenically cultivated mycobionts and photobionts by methods of UV/VIS-spectrophotometry and two types of high performance liquid chromatography (HPLC). Additionally, a set of chemical tests is presented to confirm the formation of melanic compounds in lichen and mycobiont samples. All investigated lichens reveal various sets of SLCs, except C. gyrosa where only melanin was putatively identified. Such studies will help to assess the contribution of SLCs on lichen extremotolerance, to understand the adaptation of lichens to prevalent abiotic stressors of the respective habitat, and to form a basis for interpreting recent and future astrobiological experiments. As most of the identified SLCs demonstrated a high capacity in absorbing UVR, they may also explain the high resistance of lichens towards non-terrestrial UVR.     

Evaluation of Antimicrobial Properties of Lichen Substances against Plant Pathogens

Plant pathogens pose major threats on agriculture and horticulture, causing significant economic loss worldwide. Due to the continuous and excessive use of synthetic pesticides, emergence of pesticide resistant pathogens has become more frequent. Thus, there is a growing needs for environmentally-friendly and selective antimicrobial agents with a novel mode of action, which may be used in combination with conventional pesticides to delay development of pesticide resistance. In this study, we evaluated the potentials of lichen substances as novel biopesticides against eight bacterial and twelve fungal plant pathogens that have historically caused significant phytopathological problems in South Korea. Eight lichen substances of diverse chemical origins were extracted from axenic culture or dried specimen, and further purified for comparative analysis of their antimicrobial properties. Usnic acid and vulpinic acid exhibited strong antibacterial activities against Clavibacter michiganensis subsp. michiganensis. In addition, usnic acid and vulpinic acid were highly effective in the growth inhibition of fungal pathogens, such as Diaporthe eres, D. actinidiae, and Sclerotinia sclerotiorum. Intriguingly, the growth of Rhizoctonia solani was specifically inhibited by lecanoric acid, indicating that lichen substances exhibit some degrees of selectivity to plant pathogens. These results suggested that lichen substance can be used as a selective biopesticide for controlling plant disease of agricultural and horticultural significance, minimizing possible emergence of pesticide resistant pathogens in fields.     

Ecological and biotechnological aspects of lichens

Lichens and the partners from three different kingdoms are both taxonomically and physiologically a very diverse group, which makes them interesting from both ecological and biotechnological points of view. A lichen is a mutual ecophysiological innovation in many extreme environments in which symbiosis seems to protect the partners. Lichen’s ability to grow in harsh environments can be advantageous, resulting in important ecological niches, or disadvantageous when lichens occupy and cause biodeterioration of cultural monuments. Recently, new candidate compounds for drugs, UVB protection, and antifreeze proteins for frozen foods were discovered. Lichens were also found to have potential in bioplastic degradation and prevention of desertification. Nevertheless, there is still large potential for further industrial screening and research on lichen products. Due to improved culture techniques of isolated symbionts, increased knowledge of their secondary metabolism and improved methods for solubilizing lichen metabolites, the screening and activity tests can be implemented more easily today than in the past.     

Possible functional roles of cortical depsides and medullary depsidones in the foliose lichen Hypogymnia physodes

Secondary compounds were quantified in 75 thalli of the foliose lichen Hypogymnia physodes collected in habitats along a natural shade-sun gradient from dark spruce forests to sun-exposed sea cliffs. The irradiance in all the 75 lichen sites was estimated from hemispherical photographs. The content of lichen compounds per thallus area correlated positively with irradiance level (r²=0.73), and the mean concentration increased from 6.7% in the spruce forest to 14.4% on sea cliffs. The medullary depsidones, physodic, physodalic and protocetraric acids, constituted >95% of the total pool of extractable secondary compounds, the cortical depsides, atranorin and chloratranorin, represented <5%. Both cortical compounds correlated well with direct and with diffuse radiation, whereas the three medullary compounds correlated better with diffuse than with direct radiation. Mentioned trends are consistent with a solar radiation screening hypothesis of both groups of these colourless compounds occuring as tiny crystals outside fungal hyphae. However, the UV-B protective hypothesis of the compounds was further tested in a lab experiment. Unnaturally high UV-B doses were required to significantly reduce the PSII efficiency (F_V/F_M) of symbiotic algae. Removal of the major pool of secondary compounds with acetone did not increase photobiont susceptibility to UV-B. Therefore, the main functional role of the UV-B absorbing secondary compounds in H. physodes is hardly UV-B screening. Other roles such as PAR-screening and defence against herbivores and pathogenic organisms are discussed.     

Contrasting changes in palatability following senescence of the lichenized fungi Lobaria pulmonaria and L. scrobiculata

Epiphytic lichens can contribute significantly to ecosystem nutrient input because they efficiently accumulate atmospheric mineral nutrients and, in the case of cyanolichens, also fix nitrogen. The rate at which carbon and other nutrients gained by lichens enters the ecosystem is determined by lichen litter decomposability and by invertebrate consumption of lichen litter. In turn, these processes are driven by the secondary compounds present in senesced lichens. Therefore, we explored how lichen palatability and concentrations of secondary compounds change with tissue senescence for Lobaria pulmonaria, a green-algal lichen with cyanobacterial cephalodia, and Lobaria scrobiculata, a cyanobacterial lichen. During senescence both lichens lost 38–48 % of their stictic acid chemosyndrome, while m-scrobiculin and usnic acid in L. scrobiculata remained unchanged. Snails preferred senesced rather than fresh L. pulmonaria, while senesced L. scrobiculata were avoided. This provides evidence that species with labile secondary compounds will have higher turnover rates, through consumption and decomposition, than those producing more stable secondary compounds.     

Lichen Specific Thallus Mass and Secondary Compounds Change across a Retrogressive Fire-Driven Chronosequence

In the long-term absence of major disturbances ecosystems enter a state of retrogression, which involves declining soil fertility and consequently a reduction in decomposition rates. Recent studies have looked at how plant traits such as specific leaf mass and amounts of secondary compounds respond to declining soil fertility during retrogression, but there are no comparable studies for lichen traits despite increasing recognition of the role that lichens can play in ecosystem processes. We studied a group of 30 forested islands in northern Sweden differing greatly in fire history, and collectively representing a retrogressive chronosequence, spanning 5000 years. We used this system to explore how specific thallus mass (STM) and carbon based secondary compounds (CBSCs) change in three common epiphytic lichen species (Hypogymnia phsyodes, Melanohalea olivacea and Parmelia sulcata) as soil fertility declines during this retrogression. We found that STMs of lichens increased sharply during retrogression, and for all species soil N to P ratio (which increased during retrogression) was a strong predictor of STM. When expressed per unit area, medullary CBSCs in all species and cortical CBSCs in P. sulcata increased during retrogression. Meanwhile, when expressed per unit mass, only cortical CBSCs in H. physodes responded to retrogression, and in the opposite direction. Given that lichen functional traits are likely to be important in driving ecological processes that drive nutrient and carbon cycling in the way that plant functional traits are, the changes that they undergo during retrogression could potentially be significant for the functioning of the ecosystem.     

Lichens: a promising source of antibiotic and anticancer drugs

Lichens are symbiotic associations between fungi and a photosynthetic alga and/or cyanobacteria. Lichenized fungi have been found to produce a wide array of secondary metabolites, most of which are unique to the lichenized condition. These secondary metabolites have shown an impressive range of biological activities including antibiotics, antifungal, anti-HIV, anticancer, anti-protozoan, etc. This review focuses primarily on the antibiotic and anticancer properties of lichen secondary chemicals. We have reviewed various publications related to antibiotic and anticancer drug therapies emphasizing results about specific lichens and/or lichen compounds, which microbes or cancer cells were involved and the main findings of each study. We found that crude lichen extracts and various isolated lichen compounds often demonstrate significant inhibitory activity against various pathogenic bacteria and cancer cell lines at very low concentrations. There were no studies examining the specific mechanism of action against pathogenic bacteria; however, we did find a limited number of studies where the mechanism of action against cancer cell lines had been explored. The molecular mechanism of cell death by lichen compounds includes cell cycle arrest, apoptosis, necrosis, and inhibition of angiogenesis. Although lichens are a reservoir for various biologically active compounds, only a limited number have been tested for their biological significance. There is clearly an urgent need for expanding research in this area of study, including in depth studies of those compounds which have shown promising results as well as a strong focus on identifying specific mechanisms of action and extensive clinical trials using the most promising lichen based drug therapies followed by large scale production of the best of those compounds.     

Chemical variation in the lichen genus <Emphasis Type="Italic">Letrouitia</Emphasis> (Ascomycota,Letrouitiaceae)

Secondary metabolites from 193 specimens belonging to 15 species of Letrouitia were analyzed by HPLC. Significant quantities of the anthraquinones parietin and fragilin were found in most species and occasionally minor quantities of emodin, 5-chloroemodin, 7-chloroemodin, 7-chloroteloschistin, 7-chlorofallacinal and 7-chloroparietinic acid were present. Eight previously unknown lichen substances were identified. A chemotype containing seven new dibenzofurans (8-chlorodioxocondidymic acid, 8-chlorodioxodidymic acid, 8-chloroxodidymic acid, dioxocondidymic acid, dioxodidymic acid, letrouitic acid and oxodidymic acid) was found in 3 species. In addition, a chemotype containing four unknowns (possibly chlorodepsidones) occurred in L. subvulpina. Eight different chemotypes were identified in the genus. The secondary chemistry was important for the precise identification of some species in Letrouitia. The similarity in secondary chemistry between Letrouitiaceae and Teloschistaceae is not particularly strong, as the shared compounds are also known to occur in several other lichen families.     

Lichens as a potential natural source of bioactive compounds: a review

Biological activity of material whether known in folk medicine or observed in planned screening program has been the starting point in the drug research. The general pattern is the isolation of active principles, elucidation their structures, followed by attempts for modulation of its activity potential by chemical modification. Lichens are valuable plant resources and are used as medicine, food, fodder, perfume, spice, dyes and for miscellaneous purposes throughout the world. Lichens are well known for the diversity of secondary metabolites that they produce. Compounds isolated from various lichen species have been reported to display diverse biological activities. Here we review the medicinal efficacy of lichen substances, which intends to explore the pharmaceutical potential of lichen substances.     

Do secondary substances in the thallus of a lichen promote CO2 diffusion and prevent depression of net photosynthesis at high water content?

Many lichens show seriously depressed net photosynthesis (NP) at high thallus water contents due to increased carbon dioxide diffusion resistance through blockage of diffusion pathways by water. The soil lichen Diploschistes muscorum, however, shows no depression and NP is close to maximal even at the highest thallus water content. We investigated whether lichen substances (lecanoric and diploschistesic acids) in the cortex and medulla contributed to this ability to maintain high NP. Dry thalli were extracted with water-free acetone and, after this treatment, were found to be fully viable to the extent of continued growth after replanting in the field. No differences were found in the response of NP to thallus water content between the normal and extracted thalli, in fact the response curves were often nearly identical. Thus, in this species it seems that lichen substances did not maintain the water-free diffusion pathways and some other explanation, possibly structural, needs to be sought. Received: 5 April 1997 / Accepted: 26 April 1997     

脊椎动物传播植物肉质果中的次生物质及其生态作用

在种子植物-动物的互惠关系中,植物果实成熟后需要吸引种子传播者取食果实,传播其种子至适宜萌发的生境,同时又要防御种子捕食者过度消耗种子。果实内的次生物质(如:配糖生物碱、大黄素、辣椒素)在此过程中起到重要的调控作用。依赖脊椎动物传播的肉质果中往往含有与植物茎、叶内相同的次生物质,其种类繁多,主要分为含氮化合物、酚类化合物和萜类化合物。未成熟果实内富含次生物质(如:单宁、大黄素),主要保护未成熟种子不被潜在的捕食者和食果动物取食,这些次生物质的含量通常随果实成熟而降低;其它次生物质(如:脱辅基类胡萝卜素)的含量随果实成熟而增多,可能起到吸引食果动物的作用。在对脊椎动物捕食的抵御中,果实内不同类型的次生物质促使成熟果实对所有脊椎动物都有毒性(专毒性)或者仅对种子捕食者有毒性(泛毒性)。肉质果内的次生物质对植物-食果动物相互关系的调控作用,还可以通过调节动物取食频次和数量、抑制和促进种子萌发、改变种子在肠道的滞留时间、吸引传播者等生态作用而实现。某种次生物质往往集多种生态作用于一身。目前对肉质果内次生物质与脊椎动物相互关系的探讨还不够深入。未来研究需要综合考虑植物次生物质与果实生理生化、形态学等特征对食果者的综合调控机理;次生物质在种子传播后的调控作用对植物种群或群落结构和分布格局的影响;从动植物协同进化角度探讨植物次生物质的产生、防御和吸引策略与脊椎动物对果实的选择和消费之间的关系等。开展脊椎动物传播肉质果实中次生物质的研究,对完善种子传播机制、植物繁殖和更新格局,丰富动植物相互作用、协同进化理论具有重要的意义。     

PHYSIOLOGICAL EFFECT OF LICHEN SECONDARY METABOLITES ON THE LICHEN PARASITE MARCHANDIOMYCES CORALLINUS

It has been suggested that the host specificity exhibited by some lichenicolous fungi depends on their ability to tolerate the secondary chemistry of potential host lichens. For example, the lichen parasite Marchandiomyces corallinus is able to degrade the tissues of the lichen Flavoparmelia baltimorensis irrespective of the presence or absence of endogenous phenolic compounds. In contrast, the degradation of tissues from the lichen Lasallia papulosa is suppressed when endogenous phenolics are not removed. We have investigated the physiological basis of this inhibition in order to understand more about how lichen chemistry influences host preference in lichenicolous fungi. Results showed that the secondary compounds from L. papulosa inhibit the overall growth of M. corallinus , but not the catalytic activity of its tissue-degrading polysaccharidases. This effect is different from that shown by another lichen parasite, Nectria parmeliae , where lichen compounds specifically inhibited polysaccharidase activity. Compared with the compounds of L. papulosa , the endogenous phenolics of F. baltimorensis inhibited the growth of M. corallinus substantially less and exhibited little or no inhibition of polysaccharidases. For M. corallinus , host preference appears to be associated with physiological adaptation to the chemistry of F. baltimorensis .     

Metabolic processes involved with sugar alcohol and secondary metabolite production in the hyperaccumulator lichen Diploschistes muscorum reveal its complex adaptation strategy against heavy-metal stress

The synthesis of various unique secondary metabolites by lichens is the result of mutualistic symbiotic association between the mycobiont and autotrophic photobiont. The function of these compounds and causal factors for their production are not fully understood. This paper examines the effect of heavy-metal bioaccumulation and physiological parameters related to photosynthesis and carbon metabolism on the production of lichen substances in hyperaccumulator Diploschistes muscorum. The obtained model of secondary metabolite concentrations in the thalli demonstrates that the carbon source provided by the photobiont and associated polyols produced by the mycobiont have positive impact on the production; on the contrary, the increased intracellular load of heavy metals and excessive loss of cell membrane integrity adversely affected secondary metabolite contents. Additionally, the production of secondary metabolites appears to be more dependent on intracellular metal concentrations than on soil pollution level. To compensate for metal stress, both efficient functioning of algal component and sufficient production of secondary metabolites are required. The balanced physiological functioning of mycobiont and photobiont constitutes the complex protective mechanism to alleviate the harmful effects of heavy metal stress on primary and secondary metabolism of lichens.     

Role of laccases and peroxidases in saprotrophic activities in the lichen Usnea undulata

Lichens produce various oxidoreductases including heme-containing peroxidases and the copper-containing phenol oxidases tyrosinase and laccase. Our earlier findings suggested that significant oxidoreductase activity occurs mainly in lichens from the order Peltigerales. Here we show that the non-Peltigeralean lichen Usnea can display significant activities of peroxidases and laccases. Strong evidence for the involvement of peroxidases and laccases in saprotrophic activities comes from the observation that their activities are induced by “starvation” due to prolonged dark storage, and also by treatment with soluble cellulose and lignin breakdown products. We also show that, given a quinone and chelated Fe, Usnea can produce hydroxyl radicals; these radicals contribute to the break down of carbohydrates or lignin. However, hydroxyl radical production is independent of laccase and peroxidase activity. Laccases and peroxidases are involved in other aspects of lichen biology; here we show that peroxidases, but not laccases, can break down lichen substances. Reduction in the amounts of lichen substances will reduce photoprotection, which will increase the photosynthetic capacity of thalli during winter when light intensities are low.