Low-dose ionizing radiation generates a hormetic response to modify lipid metabolism in Chlorella sorokiniana.

Algal biomass is a viable source of chemicals and metabolites for various energy, nutritional, medicinal and agricultural uses. While stresses have commonly been used to induce metabolite accumulation in microalgae in attempts to enhance high-value product yields, this is often very detrimental to growth. Therefore, understanding how to modify metabolism without deleterious consequences is highly beneficial. We demonstrate that low-doses (1-5 Gy) of ionizing radiation in the X-ray range induces a non-toxic, hormetic response in microalgae to promote metabolic activation. We identify specific radiation exposure parameters that give reproducible metabolic responses in Chlorella sorokiniana caused by transcriptional changes. This includes up-regulation of >30 lipid metabolism genes, such as genes encoding an acetyl-CoA carboxylase subunit, phosphatidic acid phosphatase, lysophosphatidic acid acyltransferase, and diacylglycerol acyltransferase. The outcome is an increased lipid yield in stationary phase cultures by 25% in just 24 hours, without any negative effects on cell viability or biomass.

Chronic toxic effects of erythromycin and its photodegradation products on microalgae Chlorella pyrenoidosa.

The photodegradation products (PDPs) of antibiotics in the aquatic environment received increasing concern, but their chronic effects on microalgae remain unclear. This study initially focused on examining the acute effects of erythromycin (ERY), then explored the chronic impacts of ERY PDPs on Chlorella pyrenoidosa. ERY of 4.0 - 32 mg/L ERY notably inhibited the cell growth and chlorophyll synthesis. The determined 96 h median effective concentration of ERY to C. pyrenoidosa was 11.78 mg/L. Higher concentrations of ERY induced more serious oxidative damage, antioxidant enzymes alleviated the oxidative stress. 6 PDPs (PDP749, PDP747, PDP719, PDP715, PDP701 and PDP557) were identified in the photodegradation process of ERY. The predicted combined toxicity of PDPs increased in the first 3 h, then decreased. Chronic exposure showed a gradual decreasing inhibition on microalgae growth and chlorophyll content. The acute effect of ERY PDPs manifested as growth stimulation, but the chronic effect manifested as growth inhibition. The malonaldehyde contents decreased with the degradation time of ERY at 7, 14 and 21 d. However, the malonaldehyde contents of ERY PDPs treatments were elevated compared to those in the control group after 21 d. Risk assessment still need to consider the potential toxicity of degradation products under long-term exposure.

The desert green algae Chlorella ohadii thrives at excessively high light intensities by exceptionally enhancing the mechanisms that protect photosynthesis from photoinhibition.

Although light is the driving force of photosynthesis, excessive light can be harmful. One of the main processes that limits photosynthesis is photoinhibition, the process of light-induced photodamage. When the absorbed light exceeds the amount that is dissipated by photosynthetic electron flow and other processes, damaging radicals are formed that mostly inactivate photosystem II (PSII). Damaged PSII must be replaced by a newly repaired complex in order to preserve full photosynthetic activity. Chlorella ohadii is a green microalga, isolated from biological desert soil crusts, that thrives under extreme high light and is highly resistant to photoinhibition. Therefore, C. ohadii is an ideal model for studying the molecular mechanisms underlying protection against photoinhibition. Comparison of the thylakoids of C. ohadii cells that were grown under low light versus extreme high light intensities found that the alga employs all three known photoinhibition protection mechanisms: (i) massive reduction of the PSII antenna size; (ii) accumulation of protective carotenoids; and (iii) very rapid repair of photodamaged reaction center proteins. This work elucidated the molecular mechanisms of photoinhibition resistance in one of the most light-tolerant photosynthetic organisms, and shows how photoinhibition protection mechanisms evolved to marginal conditions, enabling photosynthesis-dependent life in severe habitats.

Juggling Lightning: How Chlorella ohadii handles extreme energy inputs without damage.

The green alga Chlorella ohadii was isolated from a desert biological soil crust, one of the harshest environments on Earth. When grown under optimal laboratory settings it shows the fastest growth rate ever reported for a photosynthetic eukaryote and a complete resistance to photodamage even under unnaturally high light intensities. Here we examined the energy distribution along the photosynthetic pathway under four light and carbon regimes. This was performed using various methodologies such as membrane inlet mass spectrometer with stable O2 isotopes, variable fluorescence, electrochromic shift and fluorescence assessment of NADPH level, as well as the use of specific inhibitors. We show that the preceding illumination and CO2 level during growth strongly affect the energy dissipation strategies employed by the cell. For example, plastid terminal oxidase (PTOX) plays an important role in energy dissipation, particularly in high light- and low-CO2-grown cells. Of particular note is the reliance on PSII cyclic electron flow as an effective and flexible dissipation mechanism in all conditions tested. The energy management observed here may be unique to C. ohadii, as it is the only known organism to cope with such conditions. However, the strategies demonstrated may provide an insight into the processes necessary for photosynthesis under high-light conditions.

Multi-omics reveals mechanisms of total resistance to extreme illumination of a desert alga.

The unparalleled performance of Chlorella ohadii under irradiances of twice full sunlight underlines the gaps in our understanding of how the photosynthetic machinery operates, and what sets its upper functional limit. Rather than succumbing to photodamage under extreme irradiance, unique features of photosystem II function allow C. ohadii to maintain high rates of photosynthesis and growth, accompanied by major changes in composition and cellular structure. This remarkable resilience allowed us to investigate the systems response of photosynthesis and growth to extreme illumination in a metabolically active cell. Using redox proteomics, transcriptomics, metabolomics and lipidomics, we explored the cellular mechanisms that promote dissipation of excess redox energy, protein S-glutathionylation, inorganic carbon concentration, lipid and starch accumulation, and thylakoid stacking. C. ohadii possesses a readily available capacity to utilize a sudden excess of reducing power and carbon for growth and reserve formation, and post-translational redox regulation plays a pivotal role in this rapid response. Frequently the response in C. ohadii deviated from that of model species, reflecting its life history in desert sand crusts. Comparative global and case-specific analyses provided insights into the potential evolutionary role of effective reductant utilization in this extreme resistance of C. ohadii to extreme irradiation.

High light induces species specific changes in the membrane lipid composition of Chlorella.

Algae have evolved several mechanisms to adjust to changing environmental conditions. To separate from their surroundings, algal cell membranes form a hydrophobic barrier that is critical for life. Thus, it is important to maintain or adjust the physical and biochemical properties of cell membranes which are exposed to environmental factors. Especially glycerolipids of thylakoid membranes, the site of photosynthesis and photoprotection within chloroplasts, are affected by different light conditions. Since little is known about membrane lipid remodeling upon different light treatments, we examined light induced alterations in the glycerolipid composition of the two Chlorella species, C. vulgaris and C. sorokiniana, which differ strongly in their ability to cope with different light intensities. Lipidomic analysis and isotopic labeling experiments revealed differences in the composition of their galactolipid species, although both species likely utilize galactolipid precursors originated from the endoplasmic reticulum. However, in silico research of de novo sequenced genomes and ortholog mapping of proteins putatively involved in lipid metabolism showed largely conserved lipid biosynthesis pathways suggesting species specific lipid remodeling mechanisms, which possibly have an impact on the response to different light conditions.

Photoprotection mechanisms under different CO2 regimes during photosynthesis in a green alga Chlorella variabilis.

Oxygenic photosynthesis converts light energy into chemical energy via electron transport and assimilates CO2 in the Calvin-Benson cycle with the chemical energy. Thus, high light and low CO2 conditions induce the accumulation of electrons in the photosynthetic electron transport system, resulting in the formation of reactive oxygen species. To prevent the accumulation of electrons, oxygenic photosynthetic organisms have developed photoprotection mechanisms, including non-photochemical quenching (NPQ) and alternative electron flow (AEF). There are diverse molecular mechanisms underlying NPQ and AEF, and the corresponding molecular actors have been identified and characterized using a model green alga Chlamydomonas reinhardtii. In contrast, detailed information about the photoprotection mechanisms is lacking for other green algal species. In the current study, we examined the photoprotection mechanisms responsive to CO2 in the green alga Chlorella variabilis by combining the analyses of pulse-amplitude-modulated fluorescence, O2 evolution, and the steady-state and time-resolved fluorescence spectra. Under the CO2-limited condition, ΔpH-dependent NPQ occurred in photosystems I and II. Moreover, O2-dependent AEF was also induced. Under the CO2-limited condition with carbon supplementation, NPQ was relaxed and light-harvesting chlorophyll-protein complex II was isolated from both photosystems. In C. variabilis, the O2-dependent AEF and the mechanisms that instantly convert the light-harvesting functions of both photosystems may be important for maintaining efficient photosynthetic activities under various CO2 conditions.

Selenium Incorporation to Amino Acids in Chlorella Cultures Grown in Phototrophic and Heterotrophic Regimes.

Microalgae accumulate bioavailable selenium-containing amino acids (Se-AAs), and these are useful as a food supplement. While this accumulation has been studied in phototrophic algal cultures, little data exists for heterotrophic cultures. We have determined the Se-AAs content, selenium/sulfur (Se/S) substitution rates, and overall Se accumulation balance in photo- and heterotrophic Chlorella cultures. Laboratory trials revealed that heterotrophic cultures tolerate Se doses ∼8-fold higher compared to phototrophic cultures, resulting in a ∼2-3-fold higher Se-AAs content. In large-scale experiments, both cultivation regimes provided comparable Se-AAs content. Outdoor phototrophic cultures accumulated up to 400 µg g-1 of total Se-AAs and exhibited a high level of Se/S substitution (5-10%) with 30-60% organic/total Se embedded in the biomass. A slightly higher content of Se-AAs and ratio of Se/S substitution was obtained for a heterotrophic culture in pilot-scale fermentors. The data presented here shows that heterotrophic Chlorella cultures provide an alternative for Se-enriched biomass production and provides information on Se-AAs content and speciation in different cultivation regimes.

Rapid screening test to estimate temperature optima for microalgae growth using photosynthesis activity measurements.

We have worked out a rapid 1-day test based on photosynthesis measurements to estimate suitable growth temperature of microalgae cultures. To verify the proposed procedure, several microalgae-Chlorella, Nostoc, Synechocystis, Scenedesmus, and Cylindrospermum-were cultured under controlled laboratory conditions (irradiance, temperature, mixing, CO2, and nutrient supply) to find the optima of photosynthetic activity using the range between 15 and 35 °C. These activities were recorded at each temperature step after 2 h of acclimation which should be sufficient as oxygen production and the PQ cycle are regulated by fast processes. Photosynthetic activity was measured using three techniques-oxygen production/respiration, saturating pulse analysis of fluorescence quenching, and fast fluorescence induction kinetics-to estimate the temperature optima which should correspond to high growth rate. We measured all variables that might have been directly related to growth-photosynthetic oxygen evolution, maximum photochemical yield of PSII, Fv/Fm, relative electron transport rate rETRmax, and the transients Vj and Vi determined by fast fluorescence induction curves. When the temperature optima for photosynthetic activity were verified in growth tests, we found good correlation. For most of tested microalgae strains, temperature around 30 °C was found to be the most suitable at this setting. We concluded that the developed test can be used as a rapid 1-day pre-screening to estimate a suitable growth temperature of microalgae strains before they are cultured in a pilot scale.

Rapidly reversible chlorophyll fluorescence quenching induced by pulses of supersaturating light in vivo.

The saturation pulse method provides a means to distinguish between photochemical and non-photochemical quenching, based on the assumption that the former is suppressed by a saturating pulse of light (SP) and that the latter is not affected by the SP. Various types of non-photochemical quenching have been distinguished by their rates of dark relaxation in the time ranges of seconds, minutes, and hours. Here we report on a special type of non-photochemical quenching, which is rapidly induced by a pulse of high-intensity light, when PS II reaction centers are closed, and rapidly relaxes again after the pulse. This high-intensity quenching, HIQ, can be quantified by pulse-amplitude-modulation (PAM) fluorimetry (MULTI-COLOR-PAM, high sensitivity combined with high time resolution) via the quasi-instantaneous post-pulse fluorescence increase that precedes recovery of photochemical quenching in the 100-400-µs range. The HIQ amplitude increases linearly with the effective rate of quantum absorption by photosystem II, reaching about 8% of maximal fluorescence yield. It is not affected by DCMU, is stimulated by anoxic conditions, and is suppressed by energy-dependent non-photochemical quenching (NPQ). The HIQ amplitude is close to proportional to the square of maximal fluorescence yield, Fm', induced by an SP and varied by NPQ. These properties are in line with the working hypothesis of HIQ being caused by the annihilation of singlet excited chlorophyll a by triplet excited carotenoid. Significant underestimation of maximal fluorescence yield and photosystem II quantum yield in dark-acclimated samples can be avoided by use of moderate SP intensities. In physiologically healthy illuminated samples, NPQ prevents significant lowering of effective photosystem II quantum yield by HIQ, if excessive SP intensities are avoided.

Light Elicits Astaxanthin Biosynthesis and Accumulation in the Fermented Ultrahigh-Density Chlorella zofinginesis.

The growth and astaxanthin production of Chlorella zofingiensis were examined under both heterotrophic and photoautotrophic conditions, and it was found that, in comparison to the photoautotrophic mode, the heterotrophic mode led to high algal densities but attenuated intracellular astaxanthin accumulation. Following the heterotrophy-photoautotrophy transition, a considerable increase in the astaxanthin content was observed, accompanied by the upregulation of key carotenogenic genes, including phytoene synthase (PSY), ß-carotenoid hydroxylase (CHYb), ß-carotenoid ketolase 1 (BKT1), and ß-carotenoid ketolase 2 (BKT2). In contrast, the astaxanthin content and carotenogenic genes underwent an opposite change following the photoautotrophy-heterotrophy transition, suggesting the key role of light in stimulating astaxanthin biosynthesis. To improve the astaxanthin production by C. zofingiensis, a novel heterotrophy-photoinduction culture strategy without dilution was developed and evaluated. The astaxanthin content and productivity reached 2.7 mg g-1 of dry weight and 9.9 mg L-1 day-1, respectively, which were 4.0- and 2.5-fold higher than that obtained under the heterotrophic condition.

Impact of UV irradiation on Chlorella sp. damage and disinfection byproducts formation during subsequent chlorination of algal organic matter.

The frequent occurrence of algal blooms in surface water has attracted more and more attention, which caused many water quality problems, including disinfection byproducts (DBPs). Algal organic matter (AOM) including intracellular organic matter (IOM) and extracellular organic matter (EOM), was a well-known precursor to DBPs formation in drinking water. This study evaluated the effect of ultraviolet (UV) irradiation on the cell integrity, IOM release and DBPs formation during subsequent chlorination of Chlorella sp. Results showed the damage rates of algal cells increased to 40.1% after the high UV irradiation of 528 mJ/cm2, which contributed to the release of IOM. In addition, UV irradiation was effective in reducing the formation of haloacetic acids (HAAs) both in AOM and IOM, but promoted the formation of nitrogenous DBPs (N-DBPs) from AOM in subsequent chlorination. Furthermore, neutral pH exerted a positive effect on the formation of DBPs. UV irradiation decreased the bromine substitution factor (BSF) value of AOM at a high bromide level. The BSF values increased with increasing of the concentration of bromide. Moreover, more amino acids and low molecular weight precursors were produced after UV irradiation in filtered supernatant, which contributed to the formation of N-DBPs with algal chlorination. Overall, this information demonstrated pre-oxidation of UV irradiation could be used to treat the algal-rich drinking water.

Effect of light conditions on mixotrophic cultivation of green microalgae.

Current research aimed to increase mixotrophic biomass from various organic carbon sources by exploring best light conditions. Three substrates glucose, acetic acid and glycerol were studied for their effects on mixotrophic microalgae cultivation under four light conditions. Light irradiance exhibited variability in growth response and photosynthetic efficiency based on type of substrates used in mixotrophic growth. Each substrate showed variability in light requirements for their effective assimilations. From growth responses, glucose and acetic acid respectively exhibited heterotrophic and mixotrophic (better growth in light) natures. Continuous light-deficient condition was adequate for effective mixotrophic growth as well as energy saving for glucose. However, light-sufficient condition required for effective acetic acid supported mixotrophic growth. Mixotrophic benefits from glycerol and its uptake by Chlorella protothecoides was negligible in all light conditions. Investigation of heterotrophic biomass contribution by various substrates in overall mixotrophic yield, glucose offered maximum approx. 43% contribution.

Real time light intensity based carbon dioxide feeding for high cell-density microalgae cultivation and biodiesel production in a bubble column photobioreactor under outdoor natural sunlight.

Outdoor high cell-density microalgae cultivation is highly challenging due to unavailability of appropriate CO2 feeding strategy under diurnal sunlight intensities. Hence, a novel real time light based CO2 feeding strategy was firstly developed under diurnal simulated sunlight (LED) to test on Chlorella sp. in a 10 L scale bubble column photobioreactor. The strategy yielded a biomass titer of 5.12 g L-1 under simulated sunlight, far higher than existing biomass-density and pH-control based CO2 feeding strategies. In outdoor culturing, the proposed feeding strategy yielded high biomass titers of 6.8 and 9.0 g L-1 in growth-phase of two-stage and single-stage lipid induction studies respectively with same biomass productivity of 0.8 g L-1 day-1. Subsequently, two-stage lipid induction strategy of 6.8 g L-1 titer yielded biodiesel productivity of 120 g L-1 day-1, whereas single-stage strategy of 9.0 g L-1 titer was unable to induce lipid. Moreover, specific light availability affects the lipid production.

Adaptation of light-harvesting functions of unicellular green algae to different light qualities.

Oxygenic photosynthetic organisms perform photosynthesis efficiently by distributing captured light energy to photosystems (PSs) at an appropriate balance. Maintaining photosynthetic efficiency under changing light conditions requires modification of light-harvesting and energy-transfer processes. In the current study, we examined how green algae regulate their light-harvesting functions in response to different light qualities. We measured low-temperature time-resolved fluorescence spectra of unicellular green algae Chlamydomonas reinhardtii and Chlorella variabilis cells grown under different light qualities. By observing the delayed fluorescence spectra, we demonstrated that both types of green algae primarily modified the associations between light-harvesting chlorophyll protein complexes (LHCs) and PSs (PSII and PSI). Under blue light, Chlamydomonas transferred more energy from LHC to chlorophyll (Chl) located far from the PSII reaction center, while energy was transferred from LHC to PSI via different energy-transfer pathways in Chlorella. Under green light, both green algae exhibited enhanced energy transfer from LHCs to both PSs. Red light induced fluorescence quenching within PSs in Chlamydomonas and LHCs in Chlorella. In Chlorella, energy transfer from PSII to PSI appears to play an important role in balancing excitation between PSII and PSI.

High-throughput optimisation of light-driven microalgae biotechnologies.

Microalgae biotechnologies are rapidly developing into new commercial settings. Several high value products already exist on the market, and systems development is focused on cost reduction to open up future economic opportunities for food, fuel and freshwater production. Light is a key environmental driver for photosynthesis and optimising light capture is therefore critical for low cost, high efficiency systems. Here a novel high-throughput screen that simulates fluctuating light regimes in mass cultures is presented. The data was used to model photosynthetic efficiency (PEµ, mol photon-1 m2) and chlorophyll fluorescence of two green algae, Chlamydomonas reinhardtii and Chlorella sp. Response surface methodology defined the effect of three key variables: density factor (Df, 'culture density'), cycle time (tc, 'mixing rate'), and maximum incident irradiance (Imax). Both species exhibited a large rise in PEµ with decreasing Imax and a minimal effect of tc (between 3-20 s). However, the optimal Df of 0.4 for Chlamydomonas and 0.8 for Chlorella suggested strong preferences for dilute and dense cultures respectively. Chlorella had a two-fold higher optimised PEµ than Chlamydomonas, despite its higher light sensitivity. These results demonstrate species-specific light preferences within the green algae clade. Our high-throughput screen enables rapid strain selection and process optimisation.

Blue light reduces photosynthetic efficiency of cyanobacteria through an imbalance between photosystems I and II.

Several studies have described that cyanobacteria use blue light less efficiently for photosynthesis than most eukaryotic phototrophs, but comprehensive studies of this phenomenon are lacking. Here, we study the effect of blue (450 nm), orange (625 nm), and red (660 nm) light on growth of the model cyanobacterium Synechocystis sp. PCC 6803, the green alga Chlorella sorokiniana and other cyanobacteria containing phycocyanin or phycoerythrin. Our results demonstrate that specific growth rates of the cyanobacteria were similar in orange and red light, but much lower in blue light. Conversely, specific growth rates of the green alga C. sorokiniana were similar in blue and red light, but lower in orange light. Oxygen production rates of Synechocystis sp. PCC 6803 were five-fold lower in blue than in orange and red light at low light intensities but approached the same saturation level in all three colors at high light intensities. Measurements of 77 K fluorescence emission demonstrated a lower ratio of photosystem I to photosystem II (PSI:PSII ratio) and relatively more phycobilisomes associated with PSII (state 1) in blue light than in orange and red light. These results support the hypothesis that blue light, which is not absorbed by phycobilisomes, creates an imbalance between the two photosystems of cyanobacteria with an energy excess at PSI and a deficiency at the PSII-side of the photosynthetic electron transfer chain. Our results help to explain why phycobilisome-containing cyanobacteria use blue light less efficiently than species with chlorophyll-based light-harvesting antennae such as Prochlorococcus, green algae and terrestrial plants.

A comprehensive comparable study of the physiological properties of four microalgal species under different light wavelength conditions.

MAIN CONCLUSION: Microalgae treated with blue light have potential for production of human nutrition supplement and biofuel due to their higher biomass productivity and favorable fatty acid composition. Chlorella vulgaris, Chlorella pyrenoidosa, Scenedesmus quadricauda and Scenedesmus obliquus are representative green microalgae which are widely reported for algal production. In this study, we provide a systematic investigation of the biomass productivity, photosynthetic pigments, chlorophyll fluorescence and fatty acid content of the four green microalgae. The strains were grown in two primary monochromatic light wavelengths [red and blue LEDs (light emitting diode)], and in white LED conditions, respectively. Among them, blue LED light was determined as the best light for growth rate, followed by red LED and white LED. The chlorophyll generation was more sensitive to the monochromatic blue light. The polyunsaturated fatty acids (PUFAs) such as α-linolenic acid (18:3), which were perfect for human nutrition supplementation, showed high concentrations in these algae strains under blue LED. Collectively, the results indicate that the blue LED is suitable for various food, feed, and algal biofuel productions due to both biomass and fatty acid productivity.

Physiological and Ecological Aspects of Chlorella sorokiniana (Trebouxiophyceae) Under Photoautotrophic and Mixotrophic Conditions.

Mixotrophy is a metabolic strategy in which an organism is autotrophic and heterotrophic simultaneously. Considering that the aquatic environment provides several organic sources of carbon, it is probably common for microalgae to perform mixotrophy and not only photoautotrophy, but little is known about microalgae mixotrophy. The present work aimed at investigating the growth, photosynthetic activity, morphology, and biochemical composition of the microalga Chlorella sorokiniana in mixotrophic and photo-mixotrophic conditions, comparing it with photoautotrophy. The results showed pH changes after glucose addition, reaching pH 11.62 in mixotrophic and 10.47 in sequential photo-mixotrophic cultures, which limited the microalgal growth. Highest biomass was obtained in the mixotrophic culture in comparison with the sequential photo-mixotrophic one. Rapid light saturation curves showed that α (photosynthetic efficiency, 1.69) and relative electron transport rate (rETR; 565.61) were higher in the mixotrophic cultures, whereas the highest Ik (irradiance saturation, 386.68) was obtained in the photoautotrophic ones. In the sequential photo-mixotrophic cultures, photosynthetic activity varied during glucose consumption, decreasing the maximum quantum yield Fv/Fm after glucose addition, indicating change in metabolism, from photoautotrophy to mixotrophy by the microalga. The results showed that the mixotrophic cultures had higher production of chlorophyll a (6.26 mg mL-1), cell density (6.62 × 107 cell mL-1), and lipids (0.06 pg µm-3). Sequential photo-mixotrophic cultures showed the highest biovolume (360.5 µm3 cell-1) and total carbohydrates (0.026 pg µm-3). The protein concentration was 3.2 and 2.4 times higher in photoautotrophy and photo-mixotrophic growth, respectively, than in mixotrophy, but lipids were three times higher under mixotrophy. The biochemical changes we observed indicate that the microalga's plasticity in face of new environmental characteristics, such as the presence of organic carbon, can change the flow of energy through natural ecosystems.

Bleaching of biofilm-forming algae induced by UV-C treatment: a preliminary study on chlorophyll degradation and its optimization for an application on cultural heritage.

Green microalgae colonizing stone surfaces represent a major problem for the conservation of heritage monuments, since they lead to biodegradation and aesthetic issues. Previous studies in La Glacière show cave (France) have demonstrated that UV-C may have a strong effect on microalgae, thus leading to chlorophyll bleaching, which was increased when biofilms were maintained under VIS-light condition unlike to those maintained in the dark. To understand the physiological mechanisms underlying this response and in order to optimize in situ treatment, 30 kJ m-2 UV-C exposure times were applied to Chlorophyta Chlorella sp. and chlorophyll degradation kinetics were then monitored. UV-C irradiation was enough to inhibit photosynthesis and to directly kill all algal cells. Results also showed that chlorophyll a was degraded faster than chlorophyll b and that 14 h were necessary for complete degradation of all the present chlorophyll. In addition, our results highlighted the importance of visible light exposition after UV-C treatment which leading to chlorophyll bleaching. Irradiated algae cultivated in the dark were still green 5 days after treatment while cultivated samples in the light lost their green color after 14 h. An efficient UV-C treatment applicable to show caves and other heritage monuments was proposed.