Xanthoria parietina is a widespread foliose lichen growing on barks and rocks showing a broad spectrum of tolerance to air pollutants such as NOX and heavy metals, and resistance to UV-radiation because of the screening properties provided by the secondary metabolite parietin. The aim of this study was to evaluate the ability of this lichen species to survive in the following simulated space conditions, UV-radiation in N2 atmosphere and UV-radiation in vacuum. The efficiency of the photosynthetic apparatus was used as an indicator of vitality, and was expressed in terms of chlorophyll a fluorescence (FV/FM) and Normalized Difference Vegetation Index (NDVI), which were measured within 72 h from the exposure. Additionally, during the irradiation, the IR reflectance spectrum of the lichen was monitored in situ to assess changes in spectral bands. The results showed significant differences in physiological recovery trends between the treatments, highlighting that UV-radiation in vacuum causes stronger effects on FV/FM values. The IR analysis revealed several spectral band changes in the fingerprint region. The most visible variation was the 5200 cm−1 water band that disappeared in the overtone region. Nevertheless, X. parietina was able to survive UV-radiation in N2 atmosphere and in vacuum, and for this reason it may be considered a candidate for further evaluations on its survival capacity in extreme conditions.

<p>One of the main topics of astrobiology is the study of life limits in stressful environments. This field of research has the aim to understand the physiological and biochemical effects on unprotected biological samples in extreme conditions, such as space. Moreover, these studies provide indications about organisms’ adaptive plasticity under a climate change perspective, the terrestrial geological past and future scenarios, as well as extra-terrestrial habitats as Mars surface.</p> <p>The biological specimen chosen for this study was <em>Xanthoria parietina </em>(L.) Th. Fr. It is a widespread foliose lichen growing on bark and rocks which has a broad spectrum of tolerance to air pollutants such as NO<sub>X</sub> and heavy metals, and resistance to UV-radiation because of the screening properties provided by the secondary metabolism product parietin. In this study we evaluated the ability of this lichen specie to survive under simulated UV space radiation in two different extreme environments i.e., in N<sub>2</sub> atmosphere (N<sub>2</sub>) and in vacuum (10<sup>0</sup>~10<sup>-2</sup> Pa) (VAC).</p> <p>Thalli of <em>X. parietina </em>were randomly collected in a remote area of Tuscany, Italy in June and July 2020. Thalli were dehydrated for 24 h at room temperature (25°C) and stored at -18°C until treatment. Three days before the treatment, thalli were allowed to recover their normal metabolic conditions in a growth chamber at 25 °C and 70 μmol m<sup>-2</sup> s<sup>-1</sup> PAR photons. Overnight, thalli were covered with a black cotton cloth and kept moistened by spraying with distilled water.</p> <p>The simulated UV space radiation was produced using a Xe-enhanced UV lamp with a sun-like emission spectrum (wavelength range 185-2000 nm). The aforementioned atmospheric conditions (N<sub>2</sub> and VAC) were chosen to set up an extreme and dehydrating environment for the lichen. The total absorbed UV radiation dose was 1.34 MJ m<sup>-2</sup> for each exposed sample. During the irradiation, the IR reflectance spectrum of the lichen was monitored <em>in situ</em> with infrared spectroscopy to assess changes in spectral bands.</p> <p>The efficiency of the photosynthetic apparatus was assessed as indicator of vitality, and was expressed in terms of chlorophyll <em>a</em> fluorescence (F<sub>V</sub>/F<sub>M</sub>) and Normalized Difference Vegetation Index (NDVI). The examination of <em>X. parietina</em> recovery through eco-physiological analysis revealed the capacity of this lichen species to survive in extreme conditions such as those simulated in this investigation. It has been highlighted the significant difference between treatments about the photosynthetic efficiency parameters recovery trends, finding that UV-radiation in vacuum produces more intense effects on F<sub>V</sub>/F<sub>M</sub> values. After 72h, UV N<sub>2</sub> fluorescence mean values recovered up to 93% of the starting ones, while UV VAC fluorescence recovered up to 45% of the pre-exposure values. The IR analysis revealed several spectral band changes in the fingerprint region. The most visible variation was the 5200 cm<sup>-1</sup> water band, disappearing in the overtone region. This analysis suggests that the disappearance of H<sub>2</sub>O band after treatment is strictly linked to the thalli dehydration due to the atmospheric simulated conditions represented by N<sub>2</sub> insufflation and high vacuum application. Nevertheless, <em>X. parietina</em> was able to survive to UV-radiation in N<sub>2</sub> atmosphere and in vacuum, and for this reason it may be considered a candidate for further evaluations on its survival capacity in extreme conditions.</p>

Xanthoria parietina (L.) Th. Fr. is a widely spread foliose lichen showing high tolerance against UV-radiation thanks to parietin, a secondary lichen substance. We exposed samples of X. parietina under simulated Martian conditions for 30 days to explore its survivability. The lichen’s vitality was monitored via chlorophyll a fluorescence that gives an indication for active light reaction of photosynthesis, performing in situ and after-treatment analyses. Raman spectroscopy and TEM were used to evaluate carotenoid preservation and possible variations in the photobiont’s ultrastructure respectively. Significant differences in the photo-efficiency between UV irradiated samples and dark-kept samples were observed. Fluorescence values correlated with temperature and humidity day-night cycles. The photo-efficiency recovery showed that UV irradiation caused significant effects on the photosynthetic light reaction. Raman spectroscopy showed that the carotenoid signal from UV exposed samples decreased significantly after the exposure. TEM observations confirmed that UV exposed samples were the most affected by the treatment, showing chloroplastidial disorganization in photobionts’ cells. Overall, X. parietina was able to survive the simulated Mars conditions, and for this reason it may be considered as a candidate for space long-term space exposure and evaluations of the parietin photodegradability.

In this study, the combined effect of plant growth under different light quality and the application of plant-growth-promoting microbes (PGPM) was considered on spinach (Spinacia oleracea L.) to assess the influence of these factors on the photosynthetic performance. To pursue this goal, spinach plants were grown in a growth chamber at two different light quality regimes, full-spectrum white light (W) and red-blue light (RB), with (I) or without (NI) PGPM-based inoculants. Photosynthesis-light response curves (LRC) and photosynthesis-CO2 response curves (CRC) were performed for the four growth conditions (W-NI, RB-NI, W-I, and RB-I). At each step of LRC and CRC, net photosynthesis (PN), stomatal conductance (gs), Ci/Ca ratio, water use efficiency (WUEi), and fluorescence indexes were calculated. Moreover, parameters derived from the fitting of LRC, such as light-saturated net photosynthesis (PNmax), apparent light efficiency (Qpp), and dark respiration (Rd), as well as the Rubisco large subunit amount, were also determined. In not-inoculated plants, the growth under RB- regime improved PN compared to W-light because it increased stomatal conductance and favored the Rubisco synthesis. Furthermore, the RB regime also stimulates the processes of light conversion into chemical energy through chloroplasts, as indicated by the higher values of Qpp and PNmax in RB compared to W plants. On the contrary, in inoculated plants, the PN enhancement was significantly higher in W (30%) than in RB plants (17%), which showed the highest Rubisco content among all treatments. Our results indicate that the plant-growth-promoting microbes alter the photosynthetic response to light quality. This issue must be considered when PGPMs are used to improve plant growth performance in a controlled environment using artificial lighting.

<p><strong>Introduction</strong></p> <p>One of the main topics of astrobiology research is the study of life’s limits in stressful environments. The study of organisms in extreme environments might give an indication about their potential adaptive plasticity, in the view of a climate change perspective, the terrestrial geological past and future scenarios, as well as extra-terrestrial habitats such as Mars’ surface. Lichens - with their excellent adaptive abilities - represents an extremely interesting case study. Several astrobiological studies involving lichens - that are symbiotic association between a fungus and an <em>alga</em> and/or a<em>cyanobacterium</em> - proved the ability of these organisms to resist and thrive in extreme environments such as space and Mars’ surface simulated conditions [1, 2]. We have already tested the lichen species <em>Xanthoria parietina</em> (L.) Th. Fr. in simulated space conditions, that was able to survive and to reactivate after exposure [3]. <em>X. parietina</em> is a cosmopolitan foliose lichen that grows on barks and rocks [4]. This species shows high tolerance to air pollutants, heavy metals, and resistance to UV-radiation thanks to the shielding properties of the secondary metabolite parietin [5, 6]. Here we present a new study on the survival of <em>X. parietina</em> under simulated Mars conditions performed at the Mars Simulation Facility of the DLR Institute of Planetary Research in Berlin (Fig.1).</p> <p><img src="" alt="" width="257" height="451" /></p> <p>Figure 1 - Mars Simulation Facility at DLR with the opened experiment chamber.</p> <p><strong>Methods</strong></p> <p>The aim of the study was to assess the survivability of <em>Xanthoria parietina</em> under simulated Mars conditions for 30 days [7, 8]. Inside the Mars simulation chamber, eight samples (Fig.2) were exposed to the simulated atmospheric conditions of Mars of which four were fully UV-irradiated with day-night cycles (FM, Full-Mars) and the other four kept in darkness (DM, Dark-Mars). A three-gas mixture of 95% CO<sub>2</sub>, 4% N<sub>2</sub> and 1% O<sub>2</sub> was used as best approximation of Mars-like atmospheric conditions, with a constant pressure of 600Pa. Temperature and humidity were subjected to day-night cycles, reaching during daytime 15°C and 0% RH, and during night -55°C and 100% RH (Fig.3) according to Martian thermophysical conditions at mid-latitudes. UV-radiation for FM samples was simulated using a Xenon UV-lamp (spectral range 200 nm – 2200 nm) that was automatically turned on for 16 h (day) and turned off for 8 h (night) daily. The total average radiation dose for FM was 2452.32 J/cm<sup>2</sup> and the average instantaneous irradiance on the sample spots was 14,2 W/m<sup>2</sup> [9]. Four other samples (Fig.2) were kept in control conditions during the experiment, at the constant temperature of 25°C, daily wetted and 12h dark and 12h light (ca. 50 μmol m<sup>-2</sup> s<sup>-1</sup> PAR photons). Several analyses were carried out to study all the samples before, during and after the exposure to the extreme Mars conditions. In detail, this experiment was performed aiming:</p> <ul> <li>to monitor the lichen vitality through chlorophyll <em>a </em>fluorescence (F<sub>V</sub>/F<sub>M</sub>) as photosynthetic efficiency parameter, carrying out <em>in situ</em> and after treatment analyses,</li> <li>to evaluate the oxidative stress due to the extreme conditions, highlighting eventual changes in the lichen carotenoids’ Raman signatures,</li> <li>to verify eventual modifications in the infrared features (peak shifting) in the lichen FTIR reflectance spectrum possibly related to UV-photodegradation effects,</li> <li>to highlight possible variations in the lichen ultrastructure through TEM analysis.</li> </ul> <p><img src="" alt="" width="415" height="324" /></p> <p>Figure 2 - <em>Xanthoria parietina</em> samples ready for the experiment. First row (from above): full Mars samples, second row: dark Mars samples, third row: control samples.</p> <p><img src="" alt="" width="463" height="381" /></p> <p>Figure 3 - Detail of the day-night cycles of the simulated Mars conditions (temperature, red thick line; humidity, blue thin line) and fluorescence variation values for both the treatments (FM and DM).</p> <p><strong>Results</strong></p> <p>The results showed significant differences between FM and DM photosynthetic efficiency parameter during exposure to Mars environment, exhibiting F<sub>V</sub>/F<sub>M</sub> values correlated with temperature and humidity day-night cycles (Fig.3). The F<sub>V</sub>/F<sub>M</sub> recovery values showed significant differences between the treatments too, highlighting that FM conditions caused stronger effects on fluorescence values. Additional analyses show possible changes in the Raman and FTIR spectra of the irradiated samples with several features involved. Overall, <em>Xanthoria parietina</em> was able to survive to FM conditions, and for this reason it may be considered a candidate for long exposure in space and evaluations on the photodegradability of parietin in extreme conditions.</p> <p> </p> <p><strong>Reference</strong></p> <p>[1] Onofri, S., de la Torre, R., de Vera, J. P., Ott, S., Zucconi, L., Selbmann, L., Scalzi, G., Venkateswaran, K. J., Rabbow, E., Sánchez Iñigo, F. J., and Horneck, G. (2012). Survival of rock-colonizing organisms after 1.5 years in outer space. <em>Astrobiology</em>, <em>12</em>(5), 508-516.</p> <p>[2] De Vera, J. P., Möhlmann, D., Butina, F., Lorek, A., Wernecke, R., and Ott, S. (2010). Survival potential and photosynthetic activity of lichens under Mars-like conditions: a laboratory study. <em>Astrobiology</em>, <em>10</em>(2), 215-227.</p> <p>[3] Lorenz, C., Bianchi, E., Benesperi, R., Loppi, S., Papini, A., Poggiali, G., & Brucato, J. R. (2022). Survival of Xanthoria parietina in simulated space conditions: vitality assessment and spectroscopic analysis. <em>International Journal of Astrobiology</em>, 1-17.</p> <p>[4] Nimis P.L., 2016. ITALIC - The Information System on Italian Lichens. Version 5.0. University of Trieste, Dept. of Biology, (http://dryades.units.it/italic), accessed on 2022, 05, 09. for all data contained in the taxon pages, including notes, descriptions, and ecological indicator values. </p> <p>[5] Silberstein, L., Siegel, B., Siegel, S., Mukhtar, A., and Galun, M. (1996). Comparative Studies on <em>Xanthoria parietina</em>, a Pollution Resistant Lichen, and <em>Ramalina duriaei</em>, a Sensitive Species. I. Effects of Air Pollution on Physiological Processes. <em>The Lichenologist</em>, 28:355-365.</p> <p>[6] Solhaug, K. A., and Gauslaa, Y. (1996). Parietin, a photoprotective secondary product of the lichen <em>Xanthoria parietina</em>. <em>Oecologia</em>, 108:412-418.</p> <p>[7] Lorek, A., and Koncz, A. (2013). Simulation and measurement of extraterrestrial conditions for experiments on habitability with respect to Mars. In <em>Habitability of Other Planets and Satellites </em>(pp. 145-162). Springer, Dordrecht.</p> <p>[8] De Vera, J. P., Schulze-Makuch, D., Khan, A., Lorek, A., Koncz, A., Möhlmann, D., and Spohn, T. (2014). Adaptation of an Antarctic lichen to Martian niche conditions can occur within 34 days. <em>Planetary and Space Science</em>, <em>98</em>, 182-190.</p> <p>[9] Cockell, C. S., Catling, D. C., Davis, W. L., Snook, K., Kepner, R. L., Lee, P., and McKay, C. P. (2000). The ultraviolet environment of Mars: biological implications past, present, and future. <em>Icarus</em>, <em>146</em>(2), 343-359.</p>