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In bioaccumulation studies, sample devitalization through acid washing or oven drying is commonly applied to enhance the element accumulation efficiency of moss sample. Such aspect, however, has never been considered in biomonitoring surveys using lichens. In this study, the trace element accumulation performance of living (L) and dead (D) samples of the lichen Pseudevernia furfuracea was compared by a side-by-side transplanting at 40 sites in a large, mixed land use area of NE Italy for 8 weeks. Devitalization was achieved without any physico-chemical treatments, by storing lichen samples in a dark cool room for 18 months. Health status of lichens was assessed before and after the sample exposure by chlorophyll fluorescence emission. Although elemental analysis of the two exposed sample sets revealed a similar trace element pollution scenario, the content of 13 out of the 24 selected elements was higher in D samples. By expressing results as exposed-to-unexposed (EU) ratio, D samples show a higher bioaccumulation signal in 80% of transplant sites for Al, Ca, Fe, Hg, Pb and Ti. Overall, the health status of lichen samples might lead to interpretational discrepancies when EU ratio is classified according to the recently proposed bioaccumulation scale.

Kranner, Ilse/0000-0003-4959-9109; Holzinger, Andreas/0000-0002-7745-3978, PubMed: 33223597, Evidence is emerging that phytohormones represent key inter-kingdom signalling compounds supporting chemical communication between plants, fungi and bacteria. The roles of phytohormones for the lichen symbiosis are poorly understood, particularly in the process of lichenization, i.e. the key events which lead free-living microalgae and fungi to recognize each other, make physical contact and start developing a lichen thallus. Here, we studied cellular and extracellularly released phytohormones in three lichen mycobionts, Cladonia grayi, Xanthoria parietina and Tephromela atra, grown on solid medium, and the effects of indole-3-acetic acid (IAA) on their respective photobionts, Asterochloris glomerata, Trebouxia decolorans, Trebouxia sp. Using ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS) we found that mycobionts produced IAA, salicylic acid (SA) and jasmonic acid (JA). IAA represented the most abundant phytohormone produced and released by all mycobionts, whereas SA was released by X. parietina and T. atra, and JA was released by C. grayi only. With a half-life of 5.2 days, IAA degraded exponentially in solid BBM in dim light. When IAA was exogenously offered to the mycobionts' compatible photobionts at "physiological" concentrations (as released by their respective mycobionts and accumulated in the medium over seven days), the photobionts' water contents increased up to 4.4%. Treatment with IAA had no effects on the maximum quantum yield of photosystem II, dry mass, and the contents of photosynthetic pigments and alpha-tocopherol of the photobionts. The data presented may be useful for designing studies aimed at elucidating the roles of phytohormones in lichens., Austrian Science Fund (FWF)Austrian Science Fund (FWF); FWFAustrian Science Fund (FWF) [P32092-B32, I1951-B16], Open access funding provided by Austrian Science Fund (FWF).; We also thank the FWF for financial support (grants P32092-B32 to IK and I1951-B16 to AH).

Phytohormones are pivotal signaling compounds in higher plants, in which they exert their roles intracellularly, but are also released for cell-to-cell communication. In unicellular organisms, extracellularly released phytohormones can be involved in chemical crosstalk with other organisms. However, compared to higher plants, hardly any knowledge is available on the roles of phytohormones in green algae. Here, we studied phytohormone composition and extracellular release in aero-terrestrial Trebouxiophyceae. We investigated (a) which phytohormones are produced and if they are released extracellularly, and if extracellular phytohormone levels are (b) affected by environmental stimuli, and (c) differ between lichen-forming and non-lichen-forming species. Three free-living microalgae (Apatococcus lobatus, Chloroidium ellipsoideum, and Myrmecia bisecta) and three lichen-forming microalgae (Asterochloris glomerata, Trebouxia decolorans, and Trebouxia sp.) were studied. Algae were grown on solid media and the following cellular phytohormones were identified by ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS): indole-3-acetic acid (IAA), indole-3-butyric acid (IBA), abscisic acid (ABA), gibberellin A4 (GA4 ), and zeatin (ZT). Furthermore, IAA, IBA, ABA, jasmonic acid (JA), gibberellin A3 (GA3 ), and GA4 were found to be released extracellularly. IAA and ABA were released by all six species, and IAA was the most concentrated. Phytohormone release was affected by light and water availability, especially IAA in A. glomerata, Trebouxia sp., and C. ellipsoideum. No clear patterns were observed between lichen-forming and non-lichen-forming species. The results are envisaged to contribute valuable baseline information for further studies into the roles of phytohormones in microalgae.

Canopy-forming brown algae support highly productive ecosystems whose decline has been attributed to the interplay of several anthropogenic disturbances. Climate change could have disruptive effects on the biology of these species, but the role of temperature in the development of early life stages is poorly understood. The aim of this study was to assess the response of Ericaria giacconei, a winter-reproducing Southern–Mediterranean endemic species, to thermal stress by testing five temperatures (12, 15, 18, 24, and 28°C) on adults and early stages. Chlorophyll a fluorescence of adult plants was measured at 0, 24, 72, and 120 h on nine fronds in each of the three aquaria per treatment. To assess egg release, zygote settlement, and embryo growth rate, approximately 1,200 receptacles were cultured on six Petri dishes per temperature treatment, and 10 random subsections of 2 ×2 mm were examined in three Petri dishes at 0, 20, 44, and 92 h after fertilization. Adult plants showed a plastic physiological response, and thermal stress had no significant effect on PSII efficiency. Embryos fully developed only at 12 and 15°C. Mortality increased at 18 and 24°C, and no zygotes survived at 28°C. In a scenario of further increasing temperatures, the effects of warming could affect the recruitment of E. giacconei and increase its vulnerability to further stresses. These effects on the survival of early stages, which are the bottleneck for the long-term survival of the species, should be taken into account in conservation and restoration measures to maintain canopy-forming macroalgal populations and associated biodiversity and ecosystem services.

In this study, the effects of ozone (O3) on the physiology of the lichen Pseudevernia furfuracea var. furfuracea previously subjected to field stressing conditions were assessed. Samples collected in a pristine site were exposed for 6 weeks at 3 sites characterized by different pollution, e.g. elemental and PAH depositions (site RU, close to wood-burning house chimneys; site UI, close to cast-ironworks; site CK, in a semi-natural context). Afterwards, samples were transferred to controlled fumigation chambers, where they were either O3-treated for 2 weeks (250 ppb O3 for 5 h day−1, O3+ samples) or not (0 ppb O3, O3− samples). Three physiological markers (Fv/Fm, maximum quantum yield of primary photochemistry; MDA, malondialdehyde content; potassium leakage) as well as elemental and PAH concentrations were measured in matched sets of sample replicates at each experimental step. Data were explored by multivariate techniques and the effects of field exposure and fumigation were tested by generalized linear models (GLM). Detrimental effects on MDA and Fv/Fm were observed limited to samples exposed in RU and UI sites. Physiological parameters in O3-treated samples showed heterogeneous variation patterns with respect to field-exposed ones. A recovery of Fv/Fm was observed in RU- and UI-exposed samples, whereas a significant increase of MDA was highlighted limited to CK O3+ and CK O3− samples, possibly related to a “chamber effect”. Overall, the impairment caused by ozonation was limited, proving the strong O3-tolerance of our test species. Interestingly, the content of the most abundant 4-ring PAHs in RU O3+ samples, which underwent the highest field enrichment of PAHs, was significantly lower than that of matched RU O3− samples. This suggested a possible role of ozone in degrading PAHs at thallus level, with interesting interpretative repercussions in the context of transplant-based surveys aimed at evaluating PAH depositions when O3 ground levels are high.

In biomonitoring of airborne elements with native lichens, two allopatric species could be used jointly only when bioaccumulation mechanisms are the same, and differences negligible. The present study aims at (i) investigating the bioaccumulation performance for 12 elements in one-year-old lobes of Flavoparmelia caperata and Xanthoria parietina (i.e. the two lichen native biomonitors most used in the peri-Mediterranean regions) collected side-by-side on the same trees at 17 sampling sites. Both species were characterized morphologically and by X-ray diffraction analysis to evaluate specific thallus area and oxalate content, respectively. Results show that Ca and chalcophile elements (Cd, Cu, Pb and Zn) were more concentrated in F. caperata, lithophile and siderophile elements (Cr, Fe, and Ti) in X. parietina, whereas Al, Li, Mn, and Ni were accumulated in similar amounts. The outstanding content of Ca-oxalate deposits in F. caperata underlies the higher content of Ca, Cd and Pb observed in this species. On the contrary, the micro-anatomy of the upper cortex of X. parietina, which is more porous than in F. caperata, allows a higher transfer of litophile and siderophile elements. The adoption of interspecific calibration factors for biomonitoring purposes is thoroughly discussed, and the choice of using a single native species made by the most recent guidelines for biomonitoring airborne elements with lichens is fully supported.

Environmental DNA (eDNA) metabarcoding and metagenomics analyses can improve taxonomic resolution in biodiversity studies. Only recently, these techniques have been applied in aerobiology, to target bacteria, fungi and plants in airborne samples. Here, we present a nine-month aerobiological study applying eDNA metabarcoding in which we analyzed simultaneously airborne diversity and variation of fungi and plants across five locations in North and Central Italy. We correlated species composition with the ecological characteristics of the sites and the seasons. The most abundant taxa among all sites and seasons were the fungal genera Cladosporium, Alternaria, and Epicoccum and the plant genera Brassica, Corylus, Cupressus and Linum, the latter being much more variable among sites. PERMANOVA and indicator species analyses showed that the plant diversity from air samples is significantly correlated with seasons, while that of fungi varied according to the interaction between seasons and sites. The results consolidate the performance of a new eDNA metabarcoding pipeline for the simultaneous amplification and analysis of airborne plant and fungal particles. They also highlight the promising complementarity of this approach with more traditional biomonitoring frameworks and routine reports of air quality provided by environmental agencies.

Lichens can withstand extreme desiccation to water contents of ≤ 0.1 g H2O g–1 DW, and in the desiccated state are among the most extremotolerant organisms known. Desiccation-tolerant life-forms such as seeds, mosses and lichens survive ‘vitrification’, that is the transition of their cytoplasm to a ‘glassy’ state, which causes metabolism to cease. However, our understanding of the mechanisms of desiccation tolerance is hindered by poor knowledge of what reactions occur in the desiccated state. Using Flavoparmelia caperata as a model lichen, we determined at what water contents vitrification occurred upon desiccation. Molecular mobility was assessed by dynamic mechanical thermal analysis, and the de- and re-epoxidation of the xanthophyll cycle pigments (measured by HPLC) was used as a proxy to assess enzyme activity. At 20 °C vitrification occurred between 0.12–0.08 g H2O g−1 DW and enzymes were active in a ‘rubbery’ state (0.17 g H2O g−1 DW) but not in a glassy state (0.03 g H2O g−1 DW). Therefore, desiccated tissues may appear to be ‘dry’ in the conventional sense, but subtle differences in water content will have substantial consequences on the types of (bio)chemical reactions that can occur, with downstream effects on longevity in the desiccated state.

The protocols commonly applied in surveys with lichens as biomonitors of airborne trace elements require analyses of samples derived from thalli or parts of thalli grown in the last year before sampling, under the postulation that samples of the same size are of the same age. Unfortunately, the influence of ecological site-specific factors on lichen growth is still largely ignored, so that samples of the same size collected in environmentally and climatically diverse sites might actually differ in age. This work aims at quantifying the influence of climatic conditions on the radial growth rates (RaGRs) of Xanthoria parietina, one of the most popular lichen biomonitors. RaGR was monitored in seven populations distributed along an altitudinal transect of 30 km in the Classical Karst (NE Italy), from 20 to 500 m above sea level. For c. 17 months, lobe growth was measured seasonally with a digital calliper, and site-specific climatic variables were monitored by means of thermo-hygrometric sensors and implemented by meteorological data. Finally, the lobe growth of X. parietina was modelled as a function of 18 environmental variables. Results revealed that thalli of relatively dry sites had significantly lower seasonal RaGR with respect to moister ones. Considering that cumulative precipitations were equally distributed along the transect, it was concluded that RaGR of X. parietina is affected negatively by high air temperatures and positively by high relative humidity. The importance of RaGR variation in lichen bioaccumulation studies is critically discussed.

The environmental biodegradability profile of graphene related materials (GRMs) is important to know in order to predict whether these materials will accumulate in soil or will be transformed by primary decomposers. In this study, few-layer graphene (FLG) was exposed to living and devitalized axenic cultures of two white-rot basidiomycetes (Bjerkandera adusta and Phanerochaete chrysosporium) and one soil saprotrophic ascomycete (Morchella esculenta) with or without lignin, for a period of four months. Over this time, the increase of fungal biomass and presence of H2O2 and oxidizing enzymes [laccase/peroxidase and lignin peroxidase (LiP)] in growth media was assessed by gravimetric and spectrophotometric measurements, respectively. Raman spectroscopy and transmission electron microscopy (TEM) were used to compare the structure of FLG before and after incubation. All of the test fungi decreased pH in growth media and released H2O2 and laccase/peroxidase, but only basidiomycetes released LiP. Independent of growth media composition all fungi were found to be capable to oxidize FLG to a graphene oxide-like material, including M. esculenta, which released only laccase/peroxidase, i.e. the most common enzymes among primary decomposers. These findings suggest that FLG involuntarily released into terrestrial environments would likely be oxidized by soil microflora.

Heat shock treatments are an innovative and eco-friendly method to devitalize biodeteriogens of outdoor stone monuments. They consist in short (up to 6 h), mild (40–60 °C) thermal treatments applied to artificially wet surfaces, and are effective against lichens and bryophytes. Epilithic green algae, a polyphyletic and diversified group of photoautotrophs, are among the most important colonizers of stone monuments, forming conspicuous biofilms which cause chemical, physical and aesthetical damage to the substratum. Like lichens and bryophytes, they are able to face the extreme conditions of a stone surface but their resistance mechanisms are only partially known. The present study aims to test the applicability of heat shock treatments to six morphologically and phylogenetically distant green microalgae. Their survival mechanisms have been investigated in relation to photosynthesis and content of selected polyols and non-reducing sugars. Chlorophyll a fluorescence measurements and observations at the epifluorescence microscope demonstrate that the thermal treatments cause negative effects on all the species, although they do not necessarily kill the whole populations, as observed in all the lichens and bryophytes tested so far. The survival capability is discussed in relation to the production of extracellular polymeric substances and non-reducing sugars.

The interpretation of lichen bioaccumulation data is of paramount importance in environmental forensics and decision-making processes. By implementing basic ideas underlying previous interpretative scales, new dimensionless, species-independent &ldquo, bioaccumulation scales&rdquo, for native and transplanted lichens are proposed. Methodologically consistent element concentration datasets were populated with data from biomonitoring studies relying on native and transplanted lichens. The scale for native lichens was built up by analyzing the distribution of ratios between element concentration data and species-specific background concentration references (B ratios), herein provided for Flavoparmelia caperata and Xanthoria parietina (foliose lichens). The scale for transplants was built up by analyzing the distribution of ratios between element concentration in exposed and unexposed samples (EU ratio) of Evernia prunastri and Pseudevernia furfuracea (fruticose lichens). Both scales consist of five percentile-based classes, namely, &ldquo, Absence of&rdquo, &ldquo, Low&rdquo, Moderate&rdquo, High&rdquo, and &ldquo, Severe&rdquo, bioaccumulation. A comparative analysis of extant interpretative tools showed that previous ones for native lichens suffered from the obsolescence of source data, whereas the previous expert-assessed scale for transplants failed in describing noticeable element concentration variations. The new scales, based on the concept that pollution can be quantified by dimensionless ratios between experimental and benchmark values, overcome most critical points affecting the previous scales.

In bioaccumulation studies, the interpretation of pollutant contents in the target biomonitor has to be performed by assessing a deviation from an unaltered reference condition. A common strategy consists in the comparison with background element content (BEC) values, often built up by uncritically merging methodologically heterogeneous data. In this respect, the acid digestion of samples was identified as a major step affecting BEC data. Here, the analytical outcomes of two acid mixtures were compared on a set of matched paired samples of the lichen Pseudevernia furfuracea, a widely used biomonitor for which BEC values based on partial digestion were previously provided. The standard reference material BCR 482 (P. furfuracea) was used to validate analytical procedures consisting of either a HF total mineralization or an aqua regia partial one, both associated to ICP-MS multi-element analysis. In particular, the performance of the procedures was evaluated by comparing analytical results of field samples with the accuracy obtained on BCR aliquots (measured-to-expected percentage ratio). The total digestion showed a better performance for Al, As, Ba, Ca, Cd, Cu, Fe, Mn, Ni, Se, Sn, and Zn, whereas the opposite was found for Cr, Co, P, and S. Moreover, new BEC values were provided for P. furfuracea using a consolidated statistical approach, after a total sample digestion with hydrofluoric acid. The multivariate investigation of the background variability of 43 elements in 57 remote Italian sites led to the identification of geographically homogeneous areas for which BEC values are provided for use as reference in biomonitoring applications.

Melanised and non-melanised thalli of the devitalized lichen biomonitor Pseudevernia furfuracea have higher binding site concentration than mosses, adsorbing 30% more Zn2+, regardless of melanin presence. Despite the broad use of lichens as biomonitors of airborne trace elements, the surface chemistry and metal adsorption parameters of these organisms are still poorly known. The current investigation is aimed at (i) quantifying the acid-base surface properties and the first-order physical-chemical parameters of Cu2+ and Zn2+ adsorption of devitalized Pseudevernia furfuracea, a lichen commonly used in biomonitoring of airborne trace elements, and (ii) comparing the results with those available for moss biomonitors. Equilibrium constants and metal-binding site concentrations were calculated with a thermodynamic model by taking into account the presence/absence of ancillary extracellular cell wall compounds, namely melanin and acetone-soluble lichen substances. An acid–base titration experiment performed in the pH range of 3–10 showed that melanised and non-melanised P. furfuracea samples have lower pHPZC (3.53–3.99) and higher metal-binding site concentrations (0.96–1.20 mmol g−1) compared to that of the mosses investigated so far at the same experimental conditions. Melanin biosynthesis increased the content of carboxyl and phosphoryl groups and reduces that of amine/polyphenols. Cu2+ and Zn2+ adsorption was unaffected by the degree of melanisation while the removal of extracellular lichen substances slightly decreased Zn2+ adsorption. Although Cu2+ and Zn2+ adsorption parameters related to P. furfuracea surfaces were 3 times lower than in the mosses, lichen samples adsorbed the same amount of Cu2+ and 30% more Zn2+. The present study contributes in understanding the role of ancillary cell wall compounds in Cu2+ and Zn2+ adsorption in a model lichen. It also provides a first comparison between the surface physico-chemical characteristics of lichens and mosses frequently used as biomonitors of trace elements.

Products containing graphene-related materials (GRMs) are becoming quite common, raising concerns for environmental safety. GRMs have varying effects on plants, but their impact on the sexual reproduction process is largely unknown. In this study, the effects of few-layer graphene (FLG) and a similarly layered phyllosilicate, muscovite mica (MICA), were tested in vivo on the reproductive structures, i.e., pollen and stigma, of Cucurbita pepo L. ssp. pepo &lsquo, greyzini&rsquo, (summer squash, zucchini). Pollen was exposed to FLG or MICA, after careful physical-chemical characterization, at concentrations of 0.5 and 2 mg of nanomaterial (NM) per g of pollen for up to six hours. Following this, pollen viability was tested. Stigmas were exposed to FLG or MICA for three hours and then analyzed by environmental scanning electron microscopy to verify possible alterations to their surface. Stigmas were then hand-pollinated to verify the effects of the two NMs on pollen adhesion and in vivo pollen germination. FLG and MICA altered neither pollen viability nor the stigmatic surface. However, both NMs equivalently decreased pollen adhesion and in vivo germination compared with untreated stigmas. These effects deserve further attention as they could impact on production of fruits and seeds. Importantly, it was shown that FLG is as safe as a naturally occurring nanomaterial.

Drought tolerance was greater in the whole lichen than in its isolated photobiont. Cell turgor state has an influence on the functionality of photosynthetic process in lichens. Irreversible thermodynamics is widely used to describe the water relations of vascular plants. However, poikilohydrous organisms like lichens and aeroterrestrial microalgae have seldom been studied using this approach. Water relations of lichens are generally addressed without separate analysis of the mycobiont and photobiont, and only few studies have correlated changes in photosynthetic efficiency of dehydrating lichens to accurate measurements of their water potential components. We measured water potential isotherms and chlorophyll a fluorescence in the lichen Flavoparmelia caperata harvested in different seasons, as well as in its isolated photobiont, the green alga Trebouxia gelatinosa, either exposed to water stress cycles or fully hydrated. No significant seasonal trends were observed in lichen water relations parameters. Turgor loss point and osmotic potential of the whole thallus were significantly lower than those measured in the photobiont, while differences between the water stressed photobiont and controls were not significant. Dehydration-induced drop of F v/F m was correlated with turgor loss, revealing that the photosynthetic activity of lichens partly depends on their turgor level. We provided one of the first quantitative evidences of the influence that turgor status could exert on the functionality of photosynthetic processes in lichens.

The production of graphene based materials (GBMs) is steadily increasing but the effects of the possible release of GBMs in the environment are far from being understood. Graphene oxide (GO) is among the most active GBMs and it causes widely varying effects on the vegetative body of seed plants. However, nothing is known yet about its potential effects on the reproductive process. This study addresses the effects of GO on pollen germination and pollen tube elongation in the model species Nicotiana tabacum and in the non-model species Corylus avellana. In vitro germination experiments were conducted without or with GO (control and treated samples, respectively) at concentrations of 25, 50 and 100 μg mL−1. Pollen germination and tube elongation were affected at GO concentrations ≥50 μg mL−1, decreasing by 20% and 19% in N. tabacum and by 68% and 58% in C. avellana, respectively. GO did not affect the viability of N. tabacum pollen, but doubled the frequency of bent tubes. Microscopy observations of pollen tubes exposed to a cell-permeant, dual-excitation ratiometric pH indicator revealed that GO affected the intracellular pH homeostasis. Further germination experiments on C. avellana conducted by inverting the pH conditions of the control and treated (100 μg GO mL−1) samples demonstrated that the main factor influencing the pollen performance is the acidic properties of GO. This might affect the reproductive process of numerous seed plants thus being relevant from an environmental point of view.

In biomonitoring, the knowledge of background element content (BEC) values is an essential pre-requisite for the correct assessment of pollution levels. Here, we estimated the BEC values of a highly performing biomonitor, the epiphytic lichen Pseudevernia furfuracea, by means of a careful review of literature data, integrated by an extensive field survey. Methodologically homogeneous element content datasets, reflecting different exposure conditions across European and extra-European countries, were compiled and comparatively analysed. Element content in samples collected in remote areas was compared to that of potentially enriched samples, testing differences between medians for 25 elements. This analysis confirmed that the former samples were substantially unaffected by anthropogenic contributions, and their metrics were therefore proposed as a first overview at supra-national background level. We also showed that bioaccumulation studies suffer a huge methodological variability. Limited to original field data, we investigated the background variability of 43 elements in 62 remote Italian sites, characterized in GIS environment for anthropization, land use, climate and lithology at different scale resolution. The relationships between selected environmental descriptors and BEC were tested using Principal Component Regression (PCR) modelling. Elemental composition resulted significantly dependent on land use, climate and lithology. In the case of lithogenic elements, regression models correctly reproduced the lichen content throughout the country at randomly selected sites. Further descriptors should be identified only for As, Co, and V. Through a multivariate approach we also identified three geographically homogeneous macro-regions for which specific BECs were provided for use as reference in biomonitoring applications.

In this contribution, new data concerning bryophytes, fungi, and lichens of the Italian flora are presented. It includes new records and confirmations for the bryophyte genera Diplophyllum and Ptychostomum, the fungal genera Arrhenia, Gymnosporangium, and Sporidesmium and the lichen genera Arthonia, Coenogonium, Flavoplaca, Gyalolechia, Parmotrema, Peltigera, Pterygiopsis, Squamarina, Tornabea, and Waynea.

A comparative study of isolated vs. lichenized Trebouxia sp. showed that lichenization does not influence the survival capability of the alga to the photo-oxidative stress derived from prolonged desiccation. Coccoid algae in the Trebouxia genus are the most common photobionts of chlorolichens but are only sporadically found in soil or bark outside of a lichen. They all appear to be desiccation tolerant, i.e. they can survive drying to water contents of below 10 %. However, little is known about their longevity in the dry state and to which extent lichenization can influence it. Here, we studied the longevity in the dry state of the lichenized alga (LT) Trebouxia sp. in the lichen Parmotrema perlatum, in comparison with axenically grown cultures (CT) isolated from the same lichen. We report on chlorophyll fluorescence emission and reactive oxygen species (ROS) production before desiccation, after 15–45 days in the dry state under different combinations of light and air humidity and after recovery for 1 or 3 days in fully hydrated conditions. Both the CT and the LT were able to withstand desiccation under high light (120 µmol photons m−2 s−1 for 14 h per day), but upon recovery after 45 days in the dry state the performance of the CT was better than that of the LT. By contrast, the quenching of excess light energy was more efficient in the LT, at high relative humidities especially. ROS production in the LT was influenced mostly by light exposure, whereas the CT showed an oxidative burst independent of the light conditions. Although lichenization provides benefits that are essential for the survival of the photobiont in high-light habitats, Trebouxia sp. can withstand protracted periods of photo-oxidative stress even outside of a lichen thallus.

Lichens belonging to Parmeliaceae are highly diversified, but most of them share an extremely conserved morpho-chemical trait: the lower cortex is heavily melanized. The adaptive value of this character is still uncertain. Melanins are ubiquitous compounds found in most organisms since they fulfil several biological functions including defense against UV radiation, oxidizing agents, microbial stress, and metal complexation. This work aims to establish whether melanization can affect the elemental content of lichen thalli. The relative abundance of macro- (Ca, K and S) and micro- (Fe, Mn and Zn) nutrients in melanized and non-melanized pseudotissues of nine species was first evaluated by a non-destructive micro-X-ray fluorescence elemental analysis on either the upper and lower cortex, and on the internal medulla, which was artificially exposed to the mechanical removal of the lower cortex. Afterwards, the total concentration of the same elements was measured in composite samples by inductively coupled plasma atomic emission spectroscopy after acidic digestion. In order to verify whether Fe and Zn are chemically bound to the melanized pseudotissues, a sequential elution experiment was performed on two species: the two-side heavily melanized Melanelixia glabratula and the one-side lightly melanized Punctelia subrudecta. The content of Fe and Zn was higher in the melanized species than in the non-melanized ones. Species deprived of their melanized lower cortex showed a sharp decrease in Fe but not in Zn, suggesting that the melanized lower cortex is involved in Fe complexation, whereas Zn is homogeneously distributed throughout the thallus.

Removing lichen substances from dry lichen thalli using pure acetone is the least detrimental method. Measurements of properties strictly related to the photobiont, such as chlorophyllafluorescence (ChlaF), are frequently used to verify acetone toxicity but they cannot reveal possible damage accumulated at the whole thallus level. Here, measurements of ChlaF have been integrated with others concerning the status of cell membranes and photobiont population (potassium leakage, malondialdehyde and photosynthetic pigment content). Dry thalli ofFlavoparmelia caperata,Parmotrema perlatumandXanthoria parietinawere subjected to sequential acetone washings according to standard protocols. Membrane permeability was assessed before and after the washing treatment, and after a recovery period of 48 hours. Measurements of ChlaF were taken in a parallel experiment. Acetone washings increased potassium leakage in all the species from 3·9 to 6·6 times greater than the control level. After recovery, onlyP. perlatumreturned to the control level. ChlaF was affected only inF. caperata, with a 20% decrease inFv/Fmwhich had not fully recovered after 48 hours. There was neither an increase in lipid peroxidation of membranes nor a change in the photosynthetic pigment content. The sensitivity ofF. caperatato this method and the impact of the results on its future application are discussed.

The aim of this study was to verify whether it is possible to discriminate between the different pollution sources present in a mixed land use area of NE Italy on the basis of the magnetic properties and the element content of lichen transplants. Thalli of Pseudevernia furfuracea were collected in a pristine area of the South-Eastern Alps and exposed for 2 months in 40 sites located at the knots of a 700 m step grid covering ca. 40 km2 of a mosaic of agricultural, forested, industrial and urban areas. In this way, the samples could be analyzed after a defined period of time, and compared to pre-exposure conditions. The post-exposure element content and the magnetic data substantially agreed, revealing a rather modest anthropogenic impact on the territory, mostly limited to an industrial park. Since the magnetic mineralogy was homogeneous throughout the entire set of samples, with magnetite-like minerals as the main magnetic carriers, it was not possible to discriminate between PM originating from the different pollution sources. The contribution given by the industrial park could be confirmed by the multivariate analysis of the element data set. Conversely, it was possible to assess the low environmental impact of the largest local industry, a cement plant, located outside the industrial park. Notwithstanding the relatively short time of the survey, P. furfuracea was proven to be an effective accumulator for biomagnetic monitoring studies, its magnetic properties being excellent proxies for heavy metal pollution even when the anthropogenic impact on the territory is low.

Although a large body of literature exists on the use of transplanted mosses for biomonitoring of air pollution, no article has addressed so far the use and the accumulation performance of a cloned moss for this purpose. In this work, a direct comparison of metal accumulation between bags filled with a Sphagnum palustre L. clone or with native Pseudoscleropodium purum Hedw., one of the most used moss species in biomonitoring surveys, was investigated. The test was performed in sites with different atmospheric contamination levels selected in urban, industrial, agricultural and background areas of Italy and Spain. Among the eighteen elements investigated, S. palustre was significantly enriched in 10 elements (Al, Ba, Cr, Cu, Fe, Hg, Pb, Sr, V and Zn), while P. purum was enriched only in 6 elements (Al, Ba, Cu, Hg, Pb and Sr), and had a consistently lower uptake capacity than S. palustre. The clone proved to be more sensitive in terms of metal uptake and showed a better performance as a bioaccumulator, providing a higher accumulation signal and allowing a finer distinction among the different land uses and levels of pollution. The excellent uptake performance of the S. palustre clone compared to the native P. purum and its low and stable baseline elemental content, evidenced in this work, are key features for the improvement of the moss bag approach and its large scale application.

The epiphytic lichen Pseudevernia furfuracea is widely used as biomonitor of airborne trace elements and other contaminants and consists of two taxonomic varieties (var. furfuracea and var. ceratea). Here, we assessed the occurrence of inter-varietal differences in the elemental composition of paired samples of var. furfuracea and var. ceratea collected in 20 remote sites of Italian mountains. The concentration of 40 elements was measured by inductively coupled plasma mass spectroscopy, after digestion with HNO3 and aqua regia. The magnitude of inter-varietal differences compared to the effect of large-scale site-dependent environmental factors (i.e., lithological substrate, host tree species, and altitude) on overall element content was explored by multivariate analysis techniques and tested by generalized linear mixed modeling (GLMM). Further GLMMs were separately fitted for each element testing taxonomic-related variability against uncertainty associated to the analytical procedure. Inter-varietal differences were statistically significant only for Hg and P, with higher content in var. ceratea at most sites, and for Mg and Zn, showing the opposite pattern. Since the elemental composition of P. furfuracea in remote sites was mostly affected by local lithology and climatic conditions, our results confirm that lichen material for active biomonitoring should be collected in a single ecologically homogeneous remote area. We also indicate sites in the Eastern Alps where P. furfuracea showed the minimum content of most elements, which are suggested as locations to collect lichen material for transplants. Besides the context-dependency at large spatial scale, variations of elemental composition apparently related to taxonomy, could possibly be due to unequal incidence of morphological traits of the collected material. Further research is needed to clarify this issue, and how it affects bioaccumulation phenomena.

To date, there has been an ever-increasing interest in complementary air monitoring techniques, which may fill the deficiencies of air quality networks. The present work reports the results concerning five biomonitoring surveys (BSs) performed in the proximity of a waste incinerator (WI) over a 16-year period. Hg emission related to the WI activity was monitored by means of both active and passive BSs based on three photosynthetically-active biomonitors (i.e., two epiphytic lichens: Pseudevernia furfuracea and Xanthoria parietina, one vascular plant: Robinia pseudoacacia) collected or exposed before and/or after the WI installation, and after a four-month period of inactivity. Hg concentration values observed in biomonitor samples varied according to the implemented species and to the status of WI plant (active vs. inactive). Our data demonstrate that, in the same pollution scenario, P. furfuracea accumulates three times more Hg than both X. parietina and R. pseudoacacia. The results are discussed in the context of the actual European Union directives concerning air pollution monitoring and assessment, revealing that both active and passive BSs are efficient tools to provide a reliable estimation of the spatial changes of Hg concentrations in the environment.

Tropospheric ozone (O3) causes severe damage to many vascular plants but not to lichens. It was recently suggested that this may be due to their high levels of natural defences against the oxidative bursts associated to their fluctuating water content. In this study, the combined effects of watering regime (with or without a daily spray of distilled water), air relative humidity (20 ± 5 vs. 80 ± 5% RH) and O3 (250 vs. 0 ppb, 5 h day−1 for 2 weeks) were monitored in two chlorolichens with different ecology, Parmotrema perlatum and Xanthoria parietina. Modulated chlorophyll a fluorescence (Chl a F), superoxide anion radical (O2 •−) and hydrogen peroxide (H2O2) production, antioxidant content and enzyme activity of the ascorbate/glutathione cycle were measured after exposure and, for Chl a F, after 1 and 2 days of recovery. The species differed in the antioxidant profile (ascorbate was higher in X. parietina, glutathione in P. perlatum), and in the activity of ROS-scavenging enzymes, more intense in the hygrophilous P. perlatum than in the meso-xerophilous X. parietina. O3 slightly modified Chl a F parameters related to the controlled dissipation, with reduction of Fm, Fv/Fm (both species) and ETR (in P. perlatum), and increase in NPQ and qN (in X. parietina). It also influenced, particularly in P. perlatum, the content of H2O2, glutathione (GSH) and oxidized glutathione (GSSG) (but not that of O2 •− and AsA + DHA) and the activity of superoxide dismutase, ascorbate peroxidase and dehydroascorbate reductase. These parameters, however, were more heavily affected by water availability. The hypothesis that lichens are O3-tolerant thanks to the constitutive antioxidant systems, intimately related to their poikilohydric life-style, is thus confirmed.

In mountain areas of touristic interest the evaluation of the impact of human activities is crucial for ensuring long-term conservation of ecosystem biodiversity, functions and services. This study aimed at verifying the biological impact of polycyclic aromatic hydrocarbon (PAH) emissions due to traffic along the roads leading to seven passes of the Dolomites (SE Alps), which were recently declared a UNESCO World Heritage Site. Thalli of the epiphytic lichen Pseudevernia furfuracea, collected at increasing distances from the roads, were used as biomonitors. Our study revealed a gradient of decreasing PAH pollution within 300. m from the roads. Differences among passes were evident mainly for samples collected nearest to the roads, but PAH concentrations at 300. m were almost always higher than those of undisturbed reference sites, indicating that traffic PAH pollution may impact natural ecosystems and lichen diversity at relatively long distances from the emission source.

Cladina species are likely to suffer the impact of human pressure, resulting in a potential, as well as currently unknown, extinction risk for some of them. In this study, we used herbarium specimen data and literature data combined with geographic information system (GIS)-based analyses to assess the threatened status of Italian Cladina species according to IUCN criteria. A total of 485 records, reported during the period 1833–2013, were evaluated. Biological traits, habitat requirements and distribution patterns were used to infer species extinction risk. Extent of occurrence and area of occupancy have been calculated at the national scale, based on a 2km×2km cell grid. The potential threats for the taxa were assessed using a decision-support protocol in order to set conservation targets for taxa lacking population viability analyses and habitat modelling data. The species were assigned to the IUCN categories mainly using the geographical criterion B, related to species with restricted and fragmented distribution and continuous declining trend, but the species have been tested against the maximum number of criteria for which data were available and/or appropriate. This has provided an opportunity to discuss some basic aspects of the process of lichen red-listing, suggesting some methodological improvements for the mat-forming ones.

00002 ăWOS:000388157900039; International audience; This work aimed to define the molecular and chemical signature of a S. palustre clone developed in the framework of the EU-FP7 Mossclone project to improve the standardization and reliability of the moss bag technique. The molecular characterization was performed by a set of DNA molecular markers (RAPD, ISJ, PCR-RFLP, sequencing and microsatellites) to tag the clone produced within the project. Molecular characterization also provided new DNA markers that can be applied in systematic analyses of Sphagnum, and gave new insights to implement well established techniques. The elemental composition of the clone was measured by ICP-MS analysis of 54 major and trace elements, with and without commonly applied pre-exposure treatments (oven devitalization and EDTA washing). Concentrations of almost all analyzed elements were significantly lower (from 10 to 100 times) in the clone than in conspecific field moss, apart from some elements (K, Mo, P and Na) deriving from the culture medium or EDTA treatment. Oven devitalization and EDTA washing did not significantly affect the clone composition. A comparison between the elemental composition of the clone with that of naturally growing Sphagnum species proved the particularly low elemental content of the clone. Therefore, in view of a rigorously standardized moss-bag protocol for the monitoring of persistent atmospheric pollutants, the use of the S. palustre clone, a biomaterial with very low and constant element composition, and homogenous morphological characteristics is strongly recommended. (C) 2016 Elsevier Ltd. All rights reserved.

To develop an internationally standardized protocol for the moss bag technique application, the research team participating in the FP7 European project “MOSSclone” focused on the optimization of the moss bags exposure in terms of bag characteristics (shape of the bags, mesh size, weight/surface ratio), duration and height of exposure by comparing traditional moss bags to a new concept bag, “Mossphere”. In particular, the effects of each variable on the metal uptake from the air were evaluated by a systematic experimental design carried out in urban, industrial, agricultural and background areas of three European countries with oceanic, Mediterranean and continental climate. The results evidenced that the shape, the mesh size of the bags and the exposure height (in the tested ranges), did not significantly influence the uptake capacity of the transplanted moss. The aspects more affecting the element uptake were represented by the density of the moss inside the bags and the relative ratio between its weight and the surface area of the bag. We found that, the lower the density, the higher the uptake recorded. Moreover, three weeks of exposure were not enough to have a consistent uptake signal in all the environments tested, thus we suggest an exposure period not shorter than 6 weeks, which is appropriate in most situations. The above results were confirmed in all the countries and scenarios tested. The adoption of a shared exposure protocol by the research community is strongly recommended since it is a key aspect to make biomonitoring surveys directly comparable, also in view of its recognition as a monitoring method by the EU legislation.

Physicochemical deterioration processes driven by lithobiontic microcolonial fungi (MCF) and endolithic lichens (EL) are still mostly unresolved. Here, the millimetric penetration of MCF strains within silicate and carbonate lithotypes was quantified. The influence of petrographic features in determining hyphal passageways satisfies a model of physical penetration during the early stages of colonization, already described for EL. The MCF and EL secretion of iron-chelating metabolites accounts for iron mobilization in desert-varnish formation, often putatively related to fungal colonization. Increased dissolution of limestone by the model iron chelator desferrioxamine indicates the possible involvement of these MCF and EL secretes in pitting carbonates.

The hypothesis that exposure environment and land use influence element accumulation and particulate size composition in transplants of Hypnum cupressiforme has been tested using moss-bags containing oven-devitalized material. The samples were exposed for three months in ten green sites and ten roadsides in two areas with different land use (A, residential; B, residential/industrial) in the Trieste conurbation (NE Italy). Observations by SEM and EDX-ray microanalysis revealed that particle density was smaller in samples exposed in A than in B, with prevalence of particles containing Al, Ca, Fe and Si, and in good accordance with the element contents measured by acid digestion and ICP-MS. Moss-bags were generally less contaminated in green sites than in roadsides, apparently due to the different enrichment in coarse particles. In both environments, however, the majority of entrapped particles (up to 98.2%) belongs in the inhalable, small size classes (≤ PM10). The need for careful selection of the exposure sites during the phase of biomonitoring planning is discussed.

Sterile cultured isolates of lichen-forming aposymbionts have not yet been used to investigate lichen-rock interactions under controlled conditions. In this study mycobionts and photobiont of the endolithic lichens Bagliettoa baldensis and Bagliettoa marmorea were isolated and inoculated with coupons of one limestone and four marbles commonly employed in the Cultural Heritage framework. After one year of incubation, microscopic observations of polished cross-sections were performed to verify if the typical colonization patterns observed in the field may be reproduced in vitro and to evaluate the receptivity of the five lithotypes to endolithic lichens. The mycobionts of the two species developed both on the surface of and within all the lithotypes, showing different penetration pathways which depend on mineralogical and structural features and highlight different receptivity. By contrast, algae inoculated with the coupons did not penetrate them. Observations suggest that the hyphal penetration along intrinsic discontinuities of rocks is a relatively fast phenomenon when these organisms are generally considered as slow-growing. Samples from limestone outcrops and abandoned marble quarries, colonized by the same species or other representatives of Verrucariaceae, showed penetration pathways intriguingly similar to those reproduced in vitro and highlighted that lichen-driven erosion processes only increase the availability of hyphal passageways after a long-term colonization. These results show that in vitro incubation of sterile cultured lichen-forming ascomycetes with rock coupons is a practicable experimental system to investigate the lichen-rock interactions under controlled conditions and, together with analysis in situ, may support decisions on conservative treatments of historical and cultural significant stone substrata.

The intrathalline heterogeneity of chlorophyll a fluorescence (CaF) in thalli of the foliose lichen Flavoparmelia caperata was studied measuring Fo, Fm, Fv/Fm, ΔF/Fm′, NPQ and Ft in two spots of 25 mm2 positioned at ca 0.8 and 3 cm from the thallus margin. Measurements were carried out both in the field and in the laboratory using north-exposed, epiphytic thalli occurring in a deciduous wood of the Karst plateau (Trieste, NE Italy). With the exception of NPQ, all parameters were statistically higher in the marginal spots than in the central ones, suggesting that the level of photochemical activity is higher where the photobiont cells are more dense and younger. Fm and Fv/Fm of marginal spots measured in the field were statistically lower than those measured in the laboratory, probably as a consequence of the different pre-treatment conditions. The importance of selecting a specific measuring spot, particularly for biomonitoring purposes, is discussed.

Mycobiont and photobiont genetic diversity was investigated in four taxa of the Tephromela atra complex, which differ in ecology and substratum preference (from siliceous rocks, limestone to bark), and are differently interpreted by taxonomists. Phylogenetic analyses were performed using mycobiont nuclear ITS, beta tubulin and homologous polyketide synthase gene (PKS) sequences obtained from freshly collected material sampled from the Mediterranean region to the Southern Alps. The silicicolous samples from the Alps form a basal lineage of the entire complex, and despite the morphological similarity, they do not form a monophyletic group with the Mediterranean samples. No resolution was found among the calcicolous and the silicicolous taxa from Mediterranean habitats, which are traditionally segregated at variety or species level. The epiphytic taxon, although nested with the other ecotypes, splits in two well-supported lineages. Among the four taxa, Tephromela grumosa is the only morphologically, chemically and genetically distinct taxon. However, it is also nested in the large T. atra complex. Phylogenetic analysis of photobionts ITS sequences revealed that thalli from the Mediterranean region are associated with two distinct lineages of Trebouxia, but the lineages are not correlated with substrate or mycobiont phenotype. The thalli from the Alps are exclusively associated with T. simplex, suggesting a protracted isolation from the other lineages.

The efficacy of Koretrel © , a biocide used in south-eastern Asia to remove epilithic photoautotrophic organisms, was tested in the field and laboratory using lichens as target organisms. In situ observations were carried out at increasing time intervals for 6 month on 15 vegetation plots and 21 saxicolous species representative of the lichen flora of Italian monuments, selected according to their different ecology, growth-form and distribution. Change in chlorophyll fluorescence was monitored in three species for 32 day after biocide treatment. The effects of the biocide on the substratum was studied by monitoring changes in capillary water absorption and surface colorimetry of two CaCO 3 -containing rocks (Aurisina limestone and Muggia sandstone), and a grey granite. Rock crystal integrity was monitored by SEM. The results show that Koretrel © is very efficient in killing the lichens. Pronounced effects were observed in all species after 1–2 weeks, with chlorophyll fluorescence reduced to zero in 4 day in the case of Lecanora muralis and Neofuscelia loxodes and in 8 days for Caloplaca saxicola . The biocide considerably decreases capillary water absorption, and alters the surface colour of the substrata, particularly for Aurisina limestone. However, no corrosion features were observed in the rock minerals.

Trebouxia is the most common lichen-forming genus of aero-terrestrial green algae and all its species are desiccation tolerant (DT). The molecular bases of this remarkable adaptation are, however, still largely unknown. We applied a transcriptomic approach to a common member of the genus, T. gelatinosa, to investigate the alteration of gene expression occurring after dehydration and subsequent rehydration in comparison to cells kept constantly hydrated. We sequenced, de novo assembled and annotated the transcriptome of axenically cultured T. gelatinosa by using Illumina sequencing technology. We tracked the expression profiles of over 13,000 protein-coding transcripts. During the dehydration/rehydration cycle c. 92 % of the total protein-coding transcripts displayed a stable expression, suggesting that the desiccation tolerance of T. gelatinosa mostly relies on constitutive mechanisms. Dehydration and rehydration affected mainly the gene expression for components of the photosynthetic apparatus, the ROS-scavenging system, Heat Shock Proteins, aquaporins, expansins, and desiccation related proteins (DRPs), which are highly diversified in T. gelatinosa, whereas Late Embryogenesis Abundant Proteins were not affected. Only some of these phenomena were previously observed in other DT green algae, bryophytes and resurrection plants, other traits being distinctive of T. gelatinosa, and perhaps related to its symbiotic lifestyle. Finally, the phylogenetic inference extended to DRPs of other chlorophytes, embryophytes and bacteria clearly pointed out that DRPs of chlorophytes are not orthologous to those of embryophytes: some of them were likely acquired through horizontal gene transfer from extremophile bacteria which live in symbiosis within the lichen thallus.

The seasonal differences of the polycyclic aromatic hydrocarbons (PAHs) content in lichen transplants were investigated in an area of ca. 40 square kilometers in NE Italy characterized by different land use. Two sets of samples of the epiphytic lichen Pseudevernia furfuracea were collected in a pristine area of the Carnic Alps and transplanted to 40 exposure sites for a period of two months, respectively in late winter and in summer. Results revealed a pronounced difference between the two seasons in terms of PAH content and distribution patterns. After the summer exposure the PAH concentrations in the transplants were more than one order of magnitude lower than after the winter exposure (ranging from 48.22 to 272.73 ng g−1 dw and from 289.73 to 1575.85 ng g−1 dw in the summer and winter samples respectively). Also the main emission sources changed, mostly due to the drastic reduction in the emissions by wood burning for domestic heating and to the different meteorological conditions. In summer PAHs degradation was enhanced by intense UV radiation, high temperatures, and presence of ozone. The implications of these findings for the biomonitoring of PAHs pollution are addressed.

The development of isidia in thalli of Pseudevernia furfuracea from the Carnic Alps (North-eastern Italy), and the effects of these structures on CO2 gas exchanges were investigated. The ontogenetic events were studied by comparison of sections stained with different histochemical tests and SEM observations. A high cell turnover rate in both symbiotic partners is the first sign of isidium development, followed by an increased aplanosporogenesis of algae and growth of neighbouring medullary hyphae which become oriented upwards. Large nuclei and an intense cytoplasm activity characterize the mycobiont cells. The surface of very young isidia shows an irregular structure of spherical to ovoid protruding tips of perpendicular cortical hyphae, that are later organised in a pseudomeristematic area similar to that observed in the apex of growing lobes. CO2 gas exchange measurements carried out in the laboratory confirmed the high metabolic activity of isidia. At optimal water content and favourable light conditions, isolated isidia had rates of gross photosynthesis and dark respiration that were twice those of non-isidiate lobes. Isolated isidia also had a very low CO2 saturation point, probably because of their favourable surface/volume ratio, and a high light saturation, probably linked to their high content of photosynthetic pigments. The different roles played by isidia in the biology of Pseudevernia furfuracea, and particularly their rejuvenating effect on aged lobes, are discussed, and the presence of thalloconidia is briefly mentioned.

The intensity of grazing of two land snails, Cantareus aspersa and Limax sp., on three epigaeic species of Peltigera with (P. horizontalis, P. neckeri) and without (P. praetextata) secondary compounds, were studied in the field in three transects of 2×50 m along an altitudinal gradient in the surroundings of Pistoia (Tuscany, Italy). The results were confirmed by laboratory experiments and were carried also out on a further species lacking lichen compounds (P. degenii), showing that land snails definitely prefer thalli lacking lichen secondary compounds. Damage intensity is correlated to the thallus size of the lichens.

The new species Miriquidica invadens Hafellner, Obermayer & Tretiach is described and reported from Austria, Bulgaria, Italy, Switzerland, France and Spain. From a morphological point of view the species belongs to the M. griseoatra group, in which it is, so far, the only constantly lichenicolous taxon. Chemically it is characterized by the regular presence of miriquidic acid, stictic acid, and constictic acid (in variable amounts). It starts its ontogenetic life cycle as an obligate youth parasite on Sporastatia, usually S. polyspora.Morpho-anatomical treatments of other species of the Miriquidica griseoatra group occurring in Europe, based on the examination of type material, are also presented. According to the characters exhibited by the lectotype of Lecidea contigua var. griseoatra Flot., M. griseoatra (Flot.) Hertel & Rambold is the correct name for a montane species with subdivided areoles and immersed, laterally adnate apothecia with a pigmented hypothecium. Specimens exhibiting this set of characters were usually named M. obnubila auct. According to the characters observed on the lectotype of Lecidea (Biatora) obnubila Th. Fr. & Hellb., M. obnubila (Th. Fr. & Hellb.) Hertel & Rambold is a species with sessile apothecia with a hyaline hypothecium. Lecidea subplumbea Anzi does not constitute a heterotypic synonym of Lecidea griseoatra (Flot.) Schaer. but is the legitimate name at the species level for Miriquidica griseoatra sensu auct., and therefore M. subplumbea (Anzi) Cl. Roux is the correct name for that taxon. Lecidea inserena Nyl. is confirmed as a heterotypic synonym of Lecidea subplumbea Anzi. Lecidea plumbea Garov. ex A. Massal. is an earlier heterotypic synonym of Miriquidica limitata Hertel & Rambold and, therefore, the new combination M. plumbea (Garov. ex A. Massal.) Hafellner, Obermayer & Tretiach is introduced.Lectotypes are designated for Lecidea contigua var. griseoatra Flot., Lecidea inserena Nyl., Lecidea obnubila Th. Fr. & Hellb., Lecidea plumbea Garov. ex A. Massal., and Lecidea subplumbea Anzi. A key to the taxa of the M. griseoatra-group and other Miriquidica taxa with a grey thallus is provided.

The epiphytic lichenPorina pseudohibernicais described as new. It is characterized by a richly isidiate thallus, 7–8(–9)-septate ascospores and the presence of periphyses. It resemblesP. hibernicas. str., differing in the size and septation of ascospores, and in the dimensions of the terete, branched isidia. The new species was found fertile at a single site in the Carnic Alps (SE Alps, Italy), but it is also known in a sterile condition from scattered localities in the humid (sub-)montane belt of Slovenia, the Czech Republic and Ukraine, occurring on smooth bark ofFagusandCarpinus.Porina hibernicais restricted to the coastal lowlands of Atlantic Europe, and it prefers rather dry, base-rich bark ofQuercusandUlmus. The characters previously used to segregate the isidiate EuropeanPorinainto the genusZamenhofiaare also discussed.

Summary Lichens are among the best biomonitors of airborne pollutants, but surprisingly they reveal high tolerance to ozone (O3). It was recently suggested that this might be due to the high levels of natural defences against oxidative stress, related to their poikilohydric life strategy. The objective of this work is to give a thorough description of the biochemical and physiological mechanisms that are at the basis of the O3-tolerance of lichens. Chlorophyll a fluorescence (ChlaF) emission, histochemical ROS localization in the lichen thallus, and biochemical markers [enzymes and antioxidants involved in the ascorbate/glutathione (AsA/GSH) cycle; hydrogen peroxide (H2O2) and superoxide anion (O2 −)] were used to characterize the response of the epiphytic lichen Flavoparmelia caperata (L.) Hale exposed to O3 (250 ppb, 5 h d−1, 2 weeks) at different watering regimes and air relative humidity (RH) in a fumigation chamber. After two-week exposure ChlaF was affected by the watering regime but not by O3. The watering regime influenced also the superoxide dismutase activity and the production of ROS. By contrast O3 strongly influenced the AsA/GSH biochemical pathway, decreasing the reduced ascorbate (AsA) content and increasing the enzymatic activity of ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR) and glutathione reductase (GR) independently from the watering regime and the relative humidity applied. This study highlights that F. caperata can face the O3-induced oxidative stress thanks to high levels of constitutive enzymatic and non-enzymatic defences against ROS formed naturally during the dehydration–rehydration cycles to which lichens are frequently exposed.

The lichenicolous fungus “Pleospora” aquatica is described as new. It grows parasitically on thalli of Aspic&a supertegens, and is known only from the type locality, in the Tunkinski ridge (central Siberia). Is is tentatively referred to the genus PZeQspora on account of its phaeodictyospores and of the anatomy of pseudothecia. 0 1999 Adac/Editions scientifiques et

A survey aimed at studying the effects of hydrogen sulphide (H2S) on epiphytic lichen vegetation was carried out at Acquapassante (Mt. Amiata, Central Italy). In 1992, lichen vegetation was surveyed using a sampling grid often units, on 18 chestnut trees along a transect from a chimney emitting H2S to c. 200 m in the direction of the prevailing winds. A Lichen Biodiversity Index (LBI) was calculated as the sum of the frequencies of all species present within the grid. The same survey was repeated five years later. Concentration Analysis was applied to describe the data structure, and Procrustes Analysis was used to verify the congruence between the ordinations of 1992 and 1997. The statistically significant linear and non-linear regressions found between environmental variables (distance of relevés from the chimney, bark pH, lichen biomass of selected foliose and fruticose species, total sulphur content of Evernia prunastri, Hypogymnia physodes, Parmelia sulcata and Ramalina fastigiata) and the position of the relevé points on the ordination axes suggest that species distribution along the transect is related to differences in H2S tolerance. However, some crustose species (Lecanora cf. conizaeoides, L. saligna and Scolkiosporum umbrinum) should be probably excluded from the computation of the LBI for monitoring purposes, as their optimum is in the immediate vicinity of the H2S source.

Summary Six lichens [Candelariella viae-lacteae Thor & V. Wirth, Chrysothrix flavovirens Tonsberg, Psoroglaena stigonemoides (Orange) Henssen, Ramonia calcicela Canals & Gomez-Bolea, Rinodina malangica (Norman) Arnold, and Strigula calcarea Bricaud & Roux] are reported for the first time from Italy. A brief morpho-anatomical description of each species and critical notes on ecology and geographic distribution are given.

Mosses, lichens and cellulose filters were exposed for 17 weeks at four urban monitoring stations in Naples (S Italy) to assess the accumulation of airborne Al, As, Ba, Ca, Cd, Co, Cr, Cu, Fe, Hg, K, Mg, Mn, Mo, Ni, Pb, Ti, V, and Zn. In each site, the element accumulation was significantly higher in the moss Hypnum cupressiforme than in the lichen Pseudevernia furfuracea. Acid washed mosses accumulated the highest amount of trace elements, but the differences in element concentrations among the moss samples exposed after water washing and different devitalisation treatments (acid washing, oven drying and water boiling) and between the lichen samples exposed with and without the nylon bag were not statistically significant. The cellulose filters showed the lowest accumulation capability. The reciprocal ordination of sites and exposed materials showed an increasing contamination gradient (especially for Pb, Cu and Zn) from the background site to the trafficked city streets; this pattern was undetectable from PM10 data recorded by the automatic monitoring devices operating in the four exposure sites. The element profile in exposed materials did not change substantially throughout the urban area and particles of polluted urban soils seem the main source of airborne metals in Naples. Through a comprehensive evaluation of the results from this and previous studies, a protocol is suggested for the moss-bag monitoring of trace element deposition in urban environments.

Water relation parameters were measured in six congeneric lichen species with different requirements for water availability and with green algae (Peltigera aphthosa, Peltigera leucophlebia, Peltigera venosa) or cyanobacteria (Peltigera horizontalis, Peltigera praetextata, Peltigera rufescens) as main photobionts. Pressure–volume analysis was performed with a dewpoint hygrometer and integrated with anatomical analyses. The Peltigera species typical of arid environments were characterized by relatively lower osmotic potential (π0) and turgor loss point (ΨTLP), and higher values of bulk modulus of elasticity (ɛ). Both π0 and ΨTLP were correlated with the size of medullary cells, while ɛ was negatively correlated with cell dimensions. The adaptive value of low ΨTLP might reside in the capability to maintain cell turgor for longer time intervals under dry conditions. High ɛ might allow xerophilous lichens to regain cell turgor more promptly even for small amounts of water uptake, thus enlarging the cumulative period of positive carbon balance in environments with fluctuating water availability. The influence of the photobiont type is also discussed.