Phormidium ambiguum and Leptolyngbya ohadii Exopolysaccharides under Low Water Availability.

Cyanobacteria can cope with various environmental stressors, due to the excretion of exopolysaccharides (EPS). However, little is known about how the composition of these polymers may change according to water availability. This work aimed at characterizing the EPS of Phormidium ambiguum (Oscillatoriales; Oscillatoriaceae) and Leptolyngbya ohadii (Pseudanabaenales; Leptolyngbyaceae), when grown as biocrusts and biofilms, subject to water deprivation. The following EPS fractions were quantified and characterized: soluble (loosely bound, LB) and condensed (tightly bound, TB) for biocrusts, released (RPS), and sheathed in P. ambiguum and glycocalyx (G-EPS) in L. ohadii for biofilms. For both cyanobacteria upon water deprivation, glucose was the main monosaccharide present and the amount of TB-EPS resulted was significantly higher, confirming its importance in these soil-based formations. Different profiles of monosaccharides composing the EPSs were observed, as for example the higher concentration of deoxysugars observed in biocrusts compared to biofilms, demonstrating the plasticity of the cells to modify EPS composition as a response to different stresses. For both cyanobacteria, both in biofilms and biocrusts, water deprivation induced the production of simpler carbohydrates, with an increased dominance index of the composing monosaccharides. The results obtained are useful in understanding how these very relevant cyanobacterial species are sensitively modifying the EPS secreted when subject to water deprivation and could lead to consider them as suitable inoculants in degraded soils.

Assessing the Antitumor Potential of Variants of the Extracellular Carbohydrate Polymer from Synechocystis ΔsigF Mutant.

Cancer is a leading cause of death worldwide with a huge societal and economic impact. Clinically effective and less expensive anticancer agents derived from natural sources can help to overcome limitations and negative side effects of chemotherapy and radiotherapy. Previously, we showed that the extracellular carbohydrate polymer of a Synechocystis ΔsigF overproducing mutant displayed a strong antitumor activity towards several human tumor cell lines, by inducing high levels of apoptosis through p53 and caspase-3 activation. Here, the ΔsigF polymer was manipulated to obtain variants that were tested in a human melanoma (Mewo) cell line. Our results demonstrated that high molecular mass fractions were important for the polymer bioactivity, and that the reduction of the peptide content generated a variant with enhanced in vitro antitumor activity. This variant, and the original ΔsigF polymer, were further tested in vivo using the chick chorioallantoic membrane (CAM) assay. Both polymers significantly decreased xenografted CAM tumor growth and affected tumor morphology, by promoting less compact tumors, validating their antitumor potential in vivo. This work contributes with strategies for the design and testing tailored cyanobacterial extracellular polymers and further strengths the relevance of evaluating this type of polymers for biotechnological/biomedical applications.

Microalgae-Based Biorefineries: Challenges and Future Trends to Produce Carbohydrate Enriched Biomass, High-Added Value Products and Bioactive Compounds.

Microalgae have demonstrated a large potential in biotechnology as a source of various macromolecules (proteins, carbohydrates, and lipids) and high-added value products (pigments, poly-unsaturated fatty acids, peptides, exo-polysaccharides, etc.). The production of biomass at a large scale becomes more economically feasible when it is part of a biorefinery designed within the circular economy concept. Thus, the aim of this critical review is to highlight and discuss challenges and future trends related to the multi-product microalgae-based biorefineries, including both phototrophic and mixotrophic cultures treating wastewater and the recovery of biomass as a source of valuable macromolecules and high-added and low-value products (biofertilizers and biostimulants). The therapeutic properties of some microalgae-bioactive compounds are also discussed. Novel trends such as the screening of species for antimicrobial compounds, the production of bioplastics using wastewater, the circular economy strategy, and the need for more Life Cycle Assessment studies (LCA) are suggested as some of the future research lines.

Novel strategies towards efficient molecular biohydrogen production by dark fermentative mechanism: present progress and future perspective.

In the scenario of alarming increase in greenhouse and toxic gas emissions from the burning of conventional fuels, it is high time that the population drifts towards alternative fuel usage to obviate pollution. Hydrogen is an environment-friendly biofuel with high energy content. Several production methods exist to produce hydrogen, but the least energy intensive processes are the fermentative biohydrogen techniques. Dark fermentative biohydrogen production (DFBHP) is a value-added, less energy-consuming process to generate biohydrogen. In this process, biohydrogen can be produced from sugars as well as complex substrates that are generally considered as organic waste. Yet, the process is constrained by many factors such as low hydrogen yield, incomplete conversion of substrates, accumulation of volatile fatty acids which lead to the drop of the system pH resulting in hindered growth and hydrogen production by the bacteria. To circumvent these drawbacks, researchers have come up with several strategies that improve the yield of DFBHP process. These strategies can be classified as preliminary methodologies concerned with the process optimization and the latter that deals with pretreatment of substrate and seed sludge, bioaugmentation, co-culture of bacteria, supplementation of additives, bioreactor design considerations, metabolic engineering, nanotechnology, immobilization of bacteria, etc. This review sums up some of the improvement techniques that profoundly enhance the biohydrogen productivity in a DFBHP process.

In vivo anti-inflammatory and antioxidant effects of microbial polysaccharides extracted from Euganean therapeutic muds.

Therapeutic thermal mud produced by spas of the Euganean Thermal District (Italy) is used as a treatment for arthro-rheumatic diseases. Its production involves the growth of a specific microbiota embedded in a polysaccharidic matrix. Polysaccharides (Microbial-PolySaccharides, M-PS) released in the mud by the resident microorganisms were extracted and analyzed. The monosaccharidic composition analysis showed the presence of galacturonic acid, mannose, xylose, ribose and glucose and a high percentage of sulfated groups in the polymers. To assess their involvement in the therapeutic efficacy of the mud, the M-PS were tested using the model organism zebrafish (Danio rerio). The anti-inflammatory and antioxidant activities were evaluated after confirming the lack of toxic effects during development. Inflammatory state was induced chemically with copper sulfate, or through tail fin amputation procedure and UVB exposure. Recovery from inflammatory condition after exposure to M-PS was always observed with specific morphometric analyses, and further supported by qPCR. Genes linked with the inflammatory and oxidative stress response were investigated confirming the M-PS treatment's efficacy.

Rhizosheath-root system changes exopolysaccharide content but stabilizes bacterial community across contrasting seasons in a desert environment.

BACKGROUND: In hot deserts daily/seasonal fluctuations pose great challenges to the resident organisms. However, these extreme ecosystems host unique microenvironments, such as the rhizosheath-root system of desert speargrasses in which biological activities and interactions are facilitated by milder conditions and reduced fluctuations. Here, we examined the bacterial microbiota associated with this structure and its surrounding sand in the desert speargrass Stipagrostis pungens under the contrasting environmental conditions of summer and winter in the Sahara Desert. RESULTS: The belowground rhizosheath-root system has higher nutrient and humidity contents, and cooler temperatures than the surrounding sand. The plant responds to the harsh environmental conditions of the summer by increasing the abundance and diversity of extracellular polymeric substances (EPS) compared to the winter. On the contrary, the bacterial community associated with the rhizosheath-root system and its interactome remain stable and, unlike the bulk sand, are unaffected by the seasonal environmental variations. The rhizosheath-root system bacterial communities are consistently dominated by Actinobacteria and Alphaproteobacteria and form distinct bacteria communities from those of bulk sand in the two seasons. The microbiome-stabilization mediated by the plant host acts to consistently retain beneficial bacteria with multiple plant growth promoting functions, including those capable to produce EPS, which increase the sand water holding capacity ameliorating the rhizosheath micro-environment. CONCLUSIONS: Our results reveal the capability of plants in desert ecosystems to stabilize their below ground microbial community under seasonal contrasting environmental conditions, minimizing the heterogeneity of the surrounding bulk sand and contributing to the overall holobiont resilience under poly-extreme conditions.

Exopolysaccharide Features Influence Growth Success in Biocrust-forming Cyanobacteria, Moving From Liquid Culture to Sand Microcosms.

Land degradation in drylands is a drawback of the combined action of climate change and human activities. New techniques have been developed to induce artificial biocrusts formation as a tool for restoration of degraded drylands, and among them soils inoculation with cyanobacteria adapted to environmental stress. Improvement of soil properties by cyanobacteria inoculation is largely related to their ability to synthesize exopolysaccharides (EPS). However, cyanobacterial EPS features [amount, molecular weight (MW), composition] can change from one species to another or when grown in different conditions. We investigated the differences in growth and polysaccharidic matrix features among three common biocrust-forming cyanobacteria (Nostoc commune, Scytonema javanicum, and Phormidium ambiguum), when grown in liquid media and on sandy soil microcosms under optimal nutrient and water, in controlled laboratory conditions. We extracted and analyzed the released EPS (RPS) and sheath for the liquid cultures, and the more soluble or loosely-bound (LB) and the more condensed or tightly-bound (TB) soil EPS fractions for the sandy soil microcosms. In liquid culture, P. ambiguum showed the greatest growth and EPS release. In contrast, on the sandy soil, S. javanicum showed the highest growth and highest LB-EPS content. N. commune showed no relevant growth after its inoculation of the sandy soil. A difference was observed in terms of MW distribution, showing that the higher MW of the polymers produced by P. ambiguum and S. javanicum compared to the polymers produced by N. commune, could have had a positive effect on growth for the first two organisms when inoculated on the sandy soil. We also observed how both RPS and sheath fractions reflected in the composition of the soil TB-EPS fraction, indicating the role in soil stabilization of both the released and the cell attached EPS. Our results indicate that the features of the polysaccharidic matrix produced by different cyanobacteria can influence their growth success in soil. These results are of great relevance when selecting suitable candidates for large-scale cyanobacteria applications in soil restoration.

Anti-Inflammatory Activity of Exopolysaccharides from Phormidium sp. ETS05, the Most Abundant Cyanobacterium of the Therapeutic Euganean Thermal Muds, Using the Zebrafish Model.

The Euganean Thermal District (Italy) represents the oldest and largest thermal center in Europe, and its therapeutic mud is considered a unique product whose beneficial effects have been documented since Ancient Roman times. Mud properties depend on the heat and electrolytes of the thermal water, as well as on the bioactive molecules produced by its biotic component, mainly represented by cyanobacteria. The investigation of the healing effects of compounds produced by the Euganean cyanobacteria represents an important goal for scientific validation of Euganean mud therapies and for the discovering of new health beneficial biomolecules. In this work, we evaluated the therapeutic potential of exopolysaccharides (EPS) produced by Phormidium sp. ETS05, the most abundant cyanobacterium of the Euganean mud. Specifically, Phormidium EPS resulted in exerting anti-inflammatory and pro-resolution activities in chemical and injury-induced zebrafish inflammation models as demonstrated using specific transgenic zebrafish lines and morphometric and expression analyses. Moreover, in vivo and in vitro tests showed no toxicity at all for the EPS concentrations tested. The results suggest that these EPS, with their combined anti-inflammatory and pro-resolution activities, could be one of the most important therapeutic molecules present in the Euganean mud and confirm the potential of these treatments for chronic inflammatory disease recovery.

Cyanoflan: A cyanobacterial sulfated carbohydrate polymer with emulsifying properties.

The extracellular polysaccharides produced by cyanobacteria have distinctive characteristics that make them promising for applications ranging from bioremediation to biomedicine. In this study, a sulfated polysaccharide produced by a marine cyanobacterial strain and named cyanoflan was characterized in terms of morphology, chemical composition, and rheological and emulsifying properties. Cyanoflan has a 71 % carbohydrate content, with 11 % of sulfated residues, while the protein account for 4 % of dry weight. The glycosidic-substitution analysis revealed a highly branched complex chemical structure with a large number of sugar residues. The cyanoflan high molecular mass fractions (above 1 MDa) and entangled structure is consistent with its high apparent viscosity in aqueous solutions and high emulsifying activity. It showed to be a typical non-Newtonian fluid with pseudoplastic behavior. Altogether, these results confirm that cyanoflan is a versatile carbohydrate polymer that can be used in different biotechnological applications, such as emulsifying/thickening agent in food or cosmetic industries.

Characterization and antitumor activity of the extracellular carbohydrate polymer from the cyanobacterium Synechocystis ΔsigF mutant.

Cyanobacterial extracellular carbohydrate polymers are particularly attractive for biotechnological applications. Previously, we determined the monosaccharidic composition of the polymer of a Synechocystis ΔsigF overproducing mutant. Here, we further characterized this polymer, demonstrated that it is possible to recover it in high yields, and successfully use it for biomedical research. This amorphous polymer is formed by a mesh of fibrils/lamellar structures with high porosity, is constituted by high molecular mass fractions, is highly sulfated and displays low viscosity, even in highly concentrated aqueous solutions. FTIR analysis confirmed the presence of several functional groups. We demonstrated that the ΔsigF polymer has strong biological activity, decreasing the viability of melanoma, thyroid and ovary carcinoma cells by inducing high levels of apoptosis, through p53 and caspase-3 activation. Therefore, the ΔsigF Synechocystis mutant is a promising platform for the sustainable production of biological active carbohydrate polymer(s) with the desired characteristics for biomedical applications.

Increased algicidal activity of Aeromonas veronii in response to Microcystis aeruginosa: interspecies crosstalk and secondary metabolites synergism.

Toxic Microcystis spp. blooms constitute a serious threat to water quality worldwide. Aeromonas veronii was isolated from Microcystis sp. colonies collected in Lake Kinneret. Spent Aeromonas media inhibits the growth of Microcystis aeruginosa MGK isolated from Lake Kinneret. The inhibition was much stronger when Aeromonas growth medium contained spent media from MGK suggesting that Aeromonas recognized its presence and produced secondary metabolites that inhibit Microcystis growth. Fractionations of the crude extract and analyses of the active fractions identified several secondary metabolites including lumichrome in Aeromonas media. Application of lumichrome at concentrations as low as 4 nM severely inhibited Microcystis growth. Inactivation of aviH in the lumichrome biosynthetic pathway altered the lumichrome level in Aeromonas and the extent of MGK growth inhibition. Conversely, the initial lag in Aeromonas growth was significantly longer when provided with Microcystis spent media but Aeromonas was able to resume normal growth. The longer was pre-exposure to Microcystis spent media the shorter was the lag phase in Aeromonas growth indicating the presence of, and acclimation to, secondary MGK metabolite(s) the nature of which was not revealed. Our study may help to control toxic Microcystis blooms taking advantage of chemical languages used in the interspecies communication.

The role of the tyrosine kinase Wzc (Sll0923) and the phosphatase Wzb (Slr0328) in the production of extracellular polymeric substances (EPS) by Synechocystis PCC 6803.

Many cyanobacteria produce extracellular polymeric substances (EPS) mainly composed of heteropolysaccharides with unique characteristics that make them suitable for biotechnological applications. However, manipulation/optimization of EPS biosynthesis/characteristics is hindered by a poor understanding of the production pathways and the differences between bacterial species. In this work, genes putatively related to different pathways of cyanobacterial EPS polymerization, assembly, and export were targeted for deletion or truncation in the unicellular Synechocystis sp. PCC 6803. No evident phenotypic changes were observed for some mutants in genes occurring in multiple copies in Synechocystis genome, namely ∆wzy (∆sll0737), ∆wzx (∆sll5049), ∆kpsM (∆slr2107), and ∆kpsM∆wzy (∆slr2107∆sll0737), strongly suggesting functional redundancy. In contrast, Δwzc (Δsll0923) and Δwzb (Δslr0328) influenced both the amount and composition of the EPS, establishing that Wzc participates in the production of capsular (CPS) and released (RPS) polysaccharides, and Wzb affects RPS production. The structure of Wzb was solved (2.28 Å), revealing structural differences relative to other phosphatases involved in EPS production and suggesting a different substrate recognition mechanism. In addition, Wzc showed the ATPase and autokinase activities typical of bacterial tyrosine kinases. Most importantly, Wzb was able to dephosphorylate Wzc in vitro, suggesting that tyrosine phosphorylation/dephosphorylation plays a role in cyanobacterial EPS production.

Soil Type and Cyanobacteria Species Influence the Macromolecular and Chemical Characteristics of the Polysaccharidic Matrix in Induced Biocrusts.

Inoculation of soils with cyanobacteria is proposed as a sustainable biotechnological technique for restoration of degraded areas in drylands due to the important role that cyanobacteria and their exopolysaccharides (EPS) play in the environment. So far, few studies have analyzed the macromolecular and chemical characteristics of the polysaccharidic matrix in induced cyanobacterial biocrusts and the scarce existing studies have mainly focused on sandy soil textures. However, the characteristics of the cyanobacterial polysaccharidic matrix may greatly depend on soil type. The objective of this study was to examine the macromolecular distribution and monosaccharidic composition of the polysaccharidic matrix induced by inoculation of two cyanobacterial species common in arid environments, Phormidium ambiguum (non N-fixing) and Scytonema javanicum (N-fixing) in different soil types. S. javanicum promoted a higher release in the soil of the more soluble and less condensed EPS fraction (i.e., the loosely bound EPS fraction, LB-EPS), while P. ambiguum showed a higher release of the less soluble and more condensed EPS fraction (i.e., the tightly bound EPS fraction, TB-EPS). LB-EPSs were mainly composed of low MW molecules (< 50 kDa), while TB-EPSs were mainly composed of high MW molecules (1100-2000 kDa). The two EPS fractions showed a complex monosaccharidic composition (from 11 to 12 different types of monosaccharides), with glucose as the most abundant monosaccharide, in particular in the poorer soils characterized by lower organic C contents. In more C-rich soils, high abundances of galactose, mannose, and xylose were also found. Low abundance of uronic acids and hydrophobic monosaccharides, such as fucose and rhamnose, was found in the EPS extracted from the inoculated soils. Our results point to the influence of soil type on the macromolecular distribution and monosaccharide composition of the polysaccharidic matrix in induced biocrusts, which is likely to affect biocrust development and their role in soil structure and nutrient cycling in restored dryland soils.

The alternative sigma factor SigF is a key player in the control of secretion mechanisms in Synechocystis sp. PCC 6803.

Cyanobacterial alternative sigma factors are crucial players in environmental adaptation processes, which may involve bacterial responses related to maintenance of cell envelope and control of secretion pathways. Here, we show that the Group 3 alternative sigma factor F (SigF) plays a pleiotropic role in Synechocystis sp. PCC 6803 physiology, with a major impact on growth and secretion mechanisms, such as the production of extracellular polysaccharides, vesiculation and protein secretion. Although ΔsigF growth was significantly impaired, the production of released polysaccharides (RPS) increased threefold to fourfold compared with the wild-type. ΔsigF exhibits also impairment in formation of outer-membrane vesicles (OMVs) and pili, as well as several other cell envelope alterations. Similarly, the exoproteome composition of ΔsigF differs from the wild-type both in amount and type of proteins identified. Quantitative proteomics (iTRAQ) and an in silico analysis of SigF binding motifs revealed possible targets/pathways under SigF control. Besides changes in protein levels involved in secretion mechanisms, our results indicated that photosynthesis, central carbon metabolism and protein folding/degradation mechanisms are altered in ΔsigF. Overall, this work provided new evidences about the role of SigF on Synechocystis physiology and associates this regulatory element with classical and non-classical secretion pathways.

Acclimation strategy of Rhodopseudomonas palustris to high light irradiance.

The ability of Rhodopseudomonas palustris cells to rapidly acclimate to high light irradiance is an essential issue when cells are grown under sunlight. The aim of this study was to investigate the photo-acclimation process in Rhodopseudomonas palustris 42OL under different culturing conditions: (i) anaerobic (AnG), (ii) aerobic (AG), and (iii) under H2-producing (HP) conditions both at low (LL) and high light (HL) irradiances. The results obtained clearly showed that the photosynthetic unit was significantly affected by the light irradiance at which Rp. palustris 42OL was grown. The synthesis of carotenoids was affected by both illumination and culturing conditions. At LL, lycopene was the main carotenoid synthetized under all conditions tested, while at HL under HP conditions, it resulted the predominant carotenoid. Oppositely, under AnG and AG at HL, rhodovibrin was the major carotenoid detected. The increase in light intensity produced a deeper variation in light-harvesting complexes (LHC) ratio. These findings are important for understanding the ecological distribution of PNSB in natural environments, mostly characterized by high light intensities, and for its growth outdoors.

Agroindustrial residues and energy crops for the production of hydrogen and poly-ß-hydroxybutyrate via photofermentation.

The present study was aimed at assessing the biotransformation of dark fermented agroindustrial residues and energy crops for the production of hydrogen and poly-ß-hydroxybutyrate (PHB), in lab-scale photofermentation. The investigation on novel substrates for photofermentation is needed in order to enlarge the range of sustainable feedstocks. Dark fermentation effluents of ensiled maize, ensiled giant reed, ensiled olive pomace, and wheat bran were inoculated with Rhodopseudomonas palustris CGA676, a mutant strain suitable for hydrogen production in ammonium-rich media. The highest hydrogen producing performances were observed in wheat bran and maize effluents (648.6 and 320.3mLL(-1), respectively), both characterized by high initial volatile fatty acids (VFAs) concentrations. Giant reed and olive pomace effluents led to poor hydrogen production due to low initial VFAs concentrations, as the original substrates are rich in fiber. The highest PHB content was accumulated in olive pomace effluent (11.53%TS), ascribable to magnesium deficiency.

Released polysaccharides (RPS) from Cyanothece sp. CCY 0110 as biosorbent for heavy metals bioremediation: interactions between metals and RPS binding sites.

Bioremediation of heavy metals using microorganisms can be advantageous compared to conventional physicochemical methods due to the use of renewable resources and efficiencies of removal particularly cations at low concentrations. In this context, cyanobacteria/cyanobacterial extracellular polymeric substances (EPS) emerge as a valid alternative due to the anionic nature and particular composition of these polymers. In this work, various culture fractions of the unicellular cyanobacterium Cyanothece sp. CCY 0110 were employed in bioremoval assays using three of the most common heavy metal pollutants in water bodies-copper, cadmium, and lead-separately or in combined systems. Our study showed that the released polysaccharides (RPS) were the most efficient fraction, removing the metal(s) by biosorption. Therefore, this polymer was subsequently used to evaluate the interactions between the metals/RPS binding sites using SEM-EDX, ICP-OES, and FTIR. Acid and basic pretreatments applied to the polymer further improve the process efficiency, and the exposure to an alkaline solution seems to alter the RPS conformation. The differences observed in the specific metal bioremoval seem to be mainly due to the RPS organic functional groups available, mainly carboxyl and hydroxyl, than to an ion exchange mechanism. Considering that Cyanothece is a highly efficient RPS-producer and that RPS can be easily separated from the culture, immobilized or confined, this polymer can be advantageous for the establishment/improvement of heavy metal removal systems.

Energy conversion of biomass crops and agroindustrial residues by combined biohydrogen/biomethane system and anaerobic digestion.

Aim of this study was to evaluate the suitability of ensiled giant reed, ensiled maize, ensiled olive pomace, wheat bran for combined systems (CS: dark fermentation+anaerobic digestion (AD)) producing hydrogen-rich biogas (biohythane), tested in batch under basic operational conditions (mesophilic temperatures, no pH control). Substrates were also analyzed under a single stage AD batch test, in order to investigate the effects of DF on estimated energy recovery (ER) in combined systems. In CS, maize and wheat bran exhibited the highest hydrogen potential (13.8 and 18.9NLkgVS(-1)) and wheat bran the highest methane potential (243.5NLkgVS(-1)). In one-stage AD, giant reed, maize and wheat bran showed the highest methane production (239.5, 267.3 and 260.0NLkgVS(-1)). Butyrate/acetate ratio properly described the dark fermentation, correlating with hydrogen production (r=0.92). Wheat bran proved to be a promising residue for CS in terms of hydrogen/methane potential and ER.

Draft genome sequence and overview of the purple non sulfur bacterium Rhodopseudomonas palustris 42OL.

Rhodopseudomonas palustris strain 42OL was isolated in 1973 from a sugar refinery waste treatment pond. The strain has been prevalently used for hydrogen production processes using a wide variety of waste-derived substrates, and cultured both indoors and outdoors, either freely suspended or immobilized. R. palustris 42OL was suitable for many other applications and capable of growing in very different culturing conditions, revealing a wide metabolic versatility. The analysis of the genome sequence allowed to identify the metabolic pathways for hydrogen and poly-ß-hydroxy-butyrate production, and confirmed the ability of using a wide range of organic acids as substrates.

H2 production in Rhodopseudomonas palustris as a way to cope with high light intensities.

The ability of coping with the damaging effects of high light intensity represents an essential issue when purple non-sulfur bacteria (PNSB) are grown under direct sunlight for photobiological hydrogen production. This study was aimed at investigating whether H2 photo-evolution could represent, for Rhodopseudomonas palustris 42OL, a safety valve to dissipate an excess of reducing power generated under high light intensities. The physiological status of this strain was assessed under anaerobic (AnG) and aerobic (AG) growing conditions and under H2-producing (HP) conditions at low and high light intensities. The results obtained clearly showed that Fv/Fm ratio was significantly affected by the light intensity under which R. palustris 42OL cells were grown, under either AnG or AG conditions, while, under HP, it constantly remained at its highest value. The increase in light intensity significantly increased the H2 production rate, which showed a positive correlation with the maximum electron transfer rate (rETRmax). These findings are important for optimization of hydrogen production by PNSB under solar light.