Fabio Candotto Carniel

Heat Shock Treatments: A New Safe Approach against Lichen Growth on Outdoor Stone Surfaces

The control of lichen growth, particularly important in the field of stone conservation of outdoor monuments, largely depends on the use of biocides, that may be dangerous for the users, the environment and the substratum. A new, alternative approach is proposed, which makes the most of a poorly known peculiarity of poikilohydrous organisms: they are thermo-tolerant (up to 65-70 °C) when dry, but thermo-sensitive when wet. The efficacy of thermal treatments (range: 20-55 °C), in parallel to the application of three biocides, was verified in the laboratory with six epi- and endolithic lichens. Chlorophyll a fluorescence emission was checked in treated and nontreated samples of all the species, whereas histochemical observations with a dead cell stain were carried out on one of them. The feasibility of the thermal treatments in the field was verified with a seventh species. The results confirm that a 6 h treatment at 55 °C is sufficient to kill the lichens if they are kept fully hydrated. At 40 °C the organisms are damaged: in this case biocides at concentrations 10× lower than in normal applications can profitably be used. The new protocol is simple, the field equipment cheap, and the negative effects associated with standard biocide treatments are absent.

Infraspecific variability in baseline element composition of the epiphytic lichen Pseudevernia furfuracea in remote areas: implications for biomonitoring of air pollution

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.

Biomagnetic monitoring and element content of lichen transplants in a mixed land use area of NE Italy

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 2months in 40 sites located at the knots of a 700m step grid covering ca. 40km2 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.

Acetone washing for the removal of lichen substances affects membrane permeability

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 chlorophyll a fluorescence (Chl a F), are frequently used to verify acetone toxicity but they cannot reveal possible damage accumulated at the whole thallus level. Here, measurements of Chl a F have been integrated with others concerning the status of cell membranes and photobiont population (potassium leakage, malondialdehyde and photosynthetic pigment content). Dry thalli of Flavoparmelia caperata , Parmotrema perlatum and Xanthoria parietina were 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 Chl a F 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, only P. perlatum returned to the control level. Chl a F was affected only in F. caperata , with a 20% decrease in F v / F m which 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 of F. caperata to this method and the impact of the results on its future application are discussed.

Background element content of the lichen Pseudevernia furfuracea : A supra-national state of art implemented by novel field data from Italy

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.

Relation between water status and desiccation-affected genes in the lichen photobiont Trebouxia gelatinosa

The relation between water status and expression profiles of desiccation -related genes has been studied in the desiccation tolerant (DT) aeroterrestrial green microalga Trebouxia gelatinosa, a common lichen photobiont. Algal colonies were desiccated in controlled conditions and during desiccation water content (WC) and water potential (Ψ) were measured to find the turgor loss point (Ψtlp). Quantitative real-time PCR was performed to measure the expression of ten genes related to photosynthesis, antioxidant defense, expansins, heat shock proteins (HSPs), and desiccation related proteins in algal colonies collected during desiccation when still at full turgor (WC > 6 g H2O g-1 dry weight), immediately before and after Ψtlp (-4 MPa; WC ∼ 1 g H2O g-1 dry weight) and before and after complete desiccation (WC < 0.01 g H2O g-1 dry weight), quantifying the HSP70 protein levels by immunodetection. Our analysis showed that the expression of eight out of ten genes changed immediately before and after Ψtlp. Interestingly, the expression of five out of ten genes changed also before complete desiccation, i.e. between 0.2 and 0.01 g H2O g-1 dry weight. However, the HSP70 protein levels were not affected by changes in water status. The study provides new evidences of the link between the loss of turgor and the expression of genes related to the desiccation tolerance of T. gelatinosa, suggesting the former as a signal triggering inducible mechanisms.

Graphene oxide impairs the pollen performance of Nicotiana tabacum and Corylus avellana suggesting potential negative effects on the sexual reproduction of seed plants

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 cellpermeant, 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.