Bacterial tolerance strategies against lead toxicity and their relevance in bioremediation application.

Among heavy metals, lead (Pb) is a non-essential metal having a higher toxicity and without any crucial known biological functions. Being widespread, non-biodegradable and persistent in every sphere of soil, air and water, Pb is responsible for severe health and environmental issues, which need appropriate remediation measures. However, microbes inhabiting Pb-contaminated area are found to have evolved distinctive mechanisms to successfully thrive in the Pb-contaminated environment without exhibiting any negative effects on their growth and metabolism. The defensive strategies used by bacteria to ameliorate the toxic effects of lead comprise biosorption, efflux, production of metal chelators like siderophores and metallothioneins and synthesis of exopolysaccharides, extracellular sequestration and intracellular bioaccumulation. Lead remediation technologies by employing microbes may appear as potential advantageous alternatives to the conventional physical and chemical means due to specificity, suitability for applying in situ condition and feasibility to upgrade by genetic engineering. Developing strategies by designing transgenic bacterial strain having specific metal binding properties and metal chelating proteins or higher metal adsorption ability and using bacterial activity such as incorporating plant growth-promoting rhizobacteria for improved Pb resistance, exopolysaccharide and siderophores and metallothionein-mediated immobilization may prove highly effective for formulating bioremediation vis-a-vis phytoremediation strategies.

Therapeutic potential of bioactive compounds from Punica granatum extracts against aging and complicity of FOXO orthologue DAF-16 in Caenorhabditis elegans.

Some natural fruits have significant importance in improving health which provides many nutritional supplements essential to maintain proper metabolism with the age. In this study, phytochemical screening of extract (methanolic) of Punica granatum arils, outer and inner peels was confirmed by the respective spot tests. Quantification of phytochemical constituents revealed the plentiful of total phenols in the outer peels in comparison to inner peels and juice whereas total flavonoids and vitamin C are abundant in inner peel and juice, respectively. High-performance liquid chromatography, Gas chromatography along with mass spectrometry and Fourier-transform infrared spectroscopy analysis revealed the presence of compound 9, 17-octadecadienal, (Z) in the outer/inner peels. A compound N-hexadecanoic acid was also observed in the outer peels. Extracts from every section of the fruits were comprehensively evaluated for their antioxidant activity. Contrary to fruit aril juice, the extracts of outer and inner peels exhibited significant and dose-dependent in vitro antioxidant and radical-scavenging potentials. The supplementation of P. granatum extracts (PGEs) significantly enhanced the lifespan of C. elegans. The protective effect of PGEs was also observed against oxidative stress in C. elegans. Additionally, the involvement of FOXO orthologue DAF-16 dependent longevity was obtained with PGEs (outer peel and inner peel) fed TJ356 worms. Overall, the results indicate the vital role of PGEs especially the extracts of outer peels in life-saving mechanisms of C. elegans by virtue of their antioxidant asset and life-prolonging effects via daf-16 dependent Insulin signaling pathway. See also Figure 1(Fig. 1).

Cyanobacterial diversity in mat sample obtained from hypersaline desert, Rann of Kachchh.

Rann of Kachchh (RoK) is a unique geoformation, which is exposed to dynamic environmental changes such as salinity, temperature, and nutrients throughout the year. In this study, the pooled mat sample was examined for the cyanobacterial community structure using culture-dependent and culture-independent approaches. Taxonomic profiling was studied using amplicon sequencing that revealed the enrichment of Pseudanabaenales and Oscillatoriales by QIIME and MG-RAST, respectively. Other abundant orders were represented by Chroococcales, Nostocales, and unclassified cyanobacteria by both approaches. Nine cyanobacterial cultures were isolated from mat samples showing 90-98% similarities with available sequences in GenBank. The culture-dependent study suggested that mat was dominated by cyanobacterial orders such as Oscillatoriales-filamentous and Chroococcales-unicellular. Our results from the culture-dependent approach also indicated that despite high similarities in gene sequences, six cyanobacteria fall into the separate clade in the phylogenetic analysis that could be signs of evolution due to an extreme environment. Cultured isolates are correlated well with abundant taxa from amplicon sequencing. Further, protein profiling was done specifically for phycobiliproteins which will be helpful to elucidate their roles in light harvesting and energy transfer mechanism in the unique environment of RoK.

Correction to: Site, trigger, quenching mechanism and recovery of non-photochemical quenching in cyanobacteria: recent updates.

In the original publication, under the subtitle Recovery: fluorescence recovery protein (FRP), paragraph 4 the text section enclosed in quotation marks does not occur in one of the original publications cited (Sluchanko et al. 2017a, b).

Bioproduction and characterization of extracellular melanin-like pigment from industrially polluted metagenomic library equipped Escherichia coli.

To explore the potential genes from the industrially polluted Amlakhadi canal, located in Ankleshwar, Gujarat, India, its community genome was extracted and cloned into E. coli EPI300™-T1R using a fosmid vector (pCC2 FOS™) generating a library of 3,92,000 clones with average size of 40kb of DNA-insert. From this library, the clone DM1 producing brown colored melanin-like pigment was isolated and characterized. For over expression of the pigment, further sub-cloning of the clone DM1 was done. Sub-clone containing 10kb of the insert was sequenced for gene identification. The amino acids sequence of a protein 4-Hydroxyphenylpyruvate dioxygenase (HPPD), which is know to be involved in melanin biosynthesis was obtained from the gene sequence. The sequence-homology based 3D structure model of HPPD was constructed and analyzed. The physico-chemical nature of pigment was further analysed using 1H and 13C NMR, LC-MS, FTIR and UV-visible spectroscopy. The pigment was readily soluble in DMSO with an absorption maximum around 290nm. Based on the genetic and chemical characterization, the compound was confirmed as melanin-like pigment. The present results indicate that the metagenomic library from industrially polluted environment generated a microbial tool for the production of melanin-like pigment.

Site, trigger, quenching mechanism and recovery of non-photochemical quenching in cyanobacteria: recent updates.

Cyanobacteria exhibit a novel form of non-photochemical quenching (NPQ) at the level of the phycobilisome. NPQ is a process that protects photosystem II (PSII) from possible highlight-induced photo-damage. Although significant advancement has been made in understanding the NPQ, there are still some missing details. This critical review focuses on how the orange carotenoid protein (OCP) and its partner fluorescence recovery protein (FRP) control the extent of quenching. What is and what is not known about the NPQ is discussed under four subtitles; where does exactly the site of quenching lie? (site), how is the quenching being triggered? (trigger), molecular mechanism of quenching (quenching) and recovery from quenching. Finally, a recent working model of NPQ, consistent with recent findings, is been described.

Microalgal hydrogen production - A review.

Bio-hydrogen from microalgae including cyanobacteria has attracted commercial awareness due to its potential as an alternative, reliable and renewable energy source. Photosynthetic hydrogen production from microalgae can be interesting and promising options for clean energy. Advances in hydrogen-fuel-cell technology may attest an eco-friendly way of biofuel production, since, the use of H2 to generate electricity releases only water as a by-product. Progress in genetic/metabolic engineering may significantly enhance the photobiological hydrogen production from microalgae. Manipulation of competing metabolic pathways by modulating the certain key enzymes such as hydrogenase and nitrogenase may enhance the evolution of H2 from photoautotrophic cells. Moreover, biological H2 production at low operating costs is requisite for economic viability. Several photobioreactors have been developed for large-scale biomass and hydrogen production. This review highlights the recent technological progress, enzymes involved and genetic as well as metabolic engineering approaches towards sustainable hydrogen production from microalgae.

Phycoerythrin averts intracellular ROS generation and physiological functional decline in eukaryotes under oxidative stress.

In vitro antioxidant virtue and life-prolonging effect of phycoerythrin (PE; a pigment protein isolated from Phormidium sp. A09DM) have been revealed in our previous reports (Sonani et al. in Age 36:9717, 2014a; Sonani et al. in Process Biochem 49:1757-1766, 2014b). It has been hypothesized that the PE expands life span of Caenorhabditis elegans (bears large resemblance with human aging pathways) due to its antioxidant virtue. This hypothesis is tested in present study by checking the effect of PE on intracellular reactive oxygen species (ROS) generation and associated physiological deformities using mouse and human skin fibroblasts, C. elegans, and Drosophila melanogaster Oregon R + and by divulging PE's structural attributes responsible for its antioxidant asset. PE treatment displayed noteworthy decrease of 67, 48, and 77 % in ROS level in mouse fibroblast (3T3-L1), human fibroblast, and C. elegans N2, respectively, arisen under chemical-induced oxidative stress. PE treatment delayed the development of paraquat-induced Alzheimer phenotype by 14.5 % in C. elegans CL4176. Furthermore, PE improved the locomotion of D. melanogaster Oregon R + under oxidative stress with simultaneous up-regulation in super-oxide dismutase and catalase activities. The existence of 52 Glu + Asp + His + Thr residues (having metal ion sequestration capacity), 5 phycoerythrobilin chromophores (potential electron exchangers) in PE's primary structure, and significant hydrophobic patches on the surface of its α- and ß-subunits are supposed to collectively contribute in the antioxidant virtues of PE. Altogether, results support the hypothesis that it is the PE's antioxidant asset, which is responsible for its life-prolonging effect and thus could be exploited in the therapeutics of ROS-associated abnormalities including aging and neurodegeneration in eukaryotes.

Bloom Dynamics of Cyanobacteria and Their Toxins: Environmental Health Impacts and Mitigation Strategies.

Cyanobacteria are ecologically one of the most prolific groups of phototrophic prokaryotes in both marine and freshwater habitats. Both the beneficial and detrimental aspects of cyanobacteria are of considerable significance. They are important primary producers as well as an immense source of several secondary products, including an array of toxic compounds known as cyanotoxins. Abundant growth of cyanobacteria in freshwater, estuarine, and coastal ecosystems due to increased anthropogenic eutrophication and global climate change has created serious concern toward harmful bloom formation and surface water contamination all over the world. Cyanobacterial blooms and the accumulation of several cyanotoxins in water bodies pose severe ecological consequences with high risk to aquatic organisms and global public health. The proper management for mitigating the worldwide incidence of toxic cyanobacterial blooms is crucial for maintenance and sustainable development of functional ecosystems. Here, we emphasize the emerging information on the cyanobacterial bloom dynamics, toxicology of major groups of cyanotoxins, as well as a perspective and integrative approach to their management.

The high-energy radiation protectant extracellular sheath pigment scytonemin and its reduced counterpart in the cyanobacterium Scytonema sp. R77DM.

A cyanobacterial extracellular sheath pigment from Scytonema sp. R77DM was partially characterized and investigated for its increased production under abiotic factors, and UV-screening function. HPLC with PDA detection, and ion trap liquid chromatography/mass spectrometry analysis revealed the presence of a pigment scytonemin and its reduced counterpart. Ultraviolet radiation showed more stimulative effects on scytonemin production. A significant synergistic enhancement of scytonemin synthesis was observed under combined stress of heat and UV radiation. Scytonemin also exhibited efficient UV-screening function by reducing the in vivo production of reactive oxygen species (ROS) and cyclobutane thymine dimer. UV-induced formation of ROS and thymine dimer was also reduced upon exposure of cyanobacterial cells to exogenous antioxidant, ascorbic acid; however, the effect was more significant when both scytonemin and ascorbic acid were applied in combination. Moreover, the results indicate the potential role of scytonemin pigment as natural photoprotectant against high energy solar insolation.

Responses of a rice-field cyanobacterium Anabaena siamensis TISTR-8012 upon exposure to PAR and UV radiation.

The effects of PAR and UV radiation and subsequent responses of certain antioxidant enzymatic and non-enzymatic defense systems were studied in a rice field cyanobacterium Anabaena siamensis TISTR 8012. UV radiation resulted in a decline in growth accompanied by a decrease in chlorophyll a and photosynthetic efficiency. Exposure of cells to UV radiation significantly affected the differentiation of vegetative cells into heterocysts or akinetes. UV-B radiation caused the fragmentation of the cyanobacterial filaments conceivably due to the observed oxidative stress. A significant increase of reactive oxygen species in vivo and DNA strand breaks were observed in UV-B exposed cells followed by those under UV-A and PAR radiation, respectively. The UV-induced oxidative damage was alleviated due to an induction of antioxidant enzymatic/non-enzymatic defense systems. In response to UV irradiation, the studied cyanobacterium exhibited a significant increase in antioxidative enzyme activities of superoxide dismutase, catalase and peroxidase. Moreover, the cyanobacterium also synthesized some UV-absorbing/screening substances. HPLC coupled with a PDA detector revealed the presence of three compounds with UV-absorption maxima at 326, 331 and 345 nm. The induction of the biosynthesis of these UV-absorbing compounds was found under both PAR and UV radiation, thus suggesting their possible function as an active photoprotectant.

GC constituents and relative codon expressed amino acid composition in cyanobacterial phycobiliproteins.

The genomic as well as structural relationship of phycobiliproteins (PBPs) in different cyanobacterial species are determined by nucleotides as well as amino acid composition. The genomic GC constituents influence the amino acid variability and codon usage of particular subunit of PBPs. We have analyzed 11 cyanobacterial species to explore the variation of amino acids and causal relationship between GC constituents and codon usage. The study at the first, second and third levels of GC content showed relatively more amino acid variability on the levels of G3+C3 position in comparison to the first and second positions. The amino acid encoded GC rich level including G rich and C rich or both correlate the codon variability and amino acid availability. The fluctuation in amino acids such as Arg, Ala, His, Asp, Gly, Leu and Glu in α and ß subunits was observed at G1C1 position; however, fluctuation in other amino acids such as Ser, Thr, Cys and Trp was observed at G2C2 position. The coding selection pressure of amino acids such as Ala, Thr, Tyr, Asp, Gly, Ile, Leu, Asn, and Ser in α and ß subunits of PBPs was more elaborated at G3C3 position. In this study, we observed that each subunit of PBPs is codon specific for particular amino acid. These results suggest that genomic constraint linked with GC constituents selects the codon for particular amino acids and furthermore, the codon level study may be a novel approach to explore many problems associated with genomics and proteomics of cyanobacteria.

Analysis of UV-absorbing photoprotectant mycosporine-like amino acid (MAA) in the cyanobacterium Arthrospira sp. CU2556.

Mycosporine-like amino acids (MAAs) are ecologically important biomolecules with great photoprotective potential. The present study aimed to investigate the biosynthesis of MAAs in the cyanobacterium Arthrospira sp. CU2556. High-performance liquid chromatography (HPLC) with photodiode-array detection studies revealed the presence of a UV-absorbing compound with an absorption maximum at 310 nm. Based on its UV absorption spectrum and ion trap liquid chromatography/mass spectrometry (LC/MS) analysis, the compound was identified as a primary MAA mycosporine-glycine (m/z: 246). To the best of our knowledge this is the first report on the occurrence of MAA mycosporine-glycine (M-Gly) in Arthrospira strains studied so far. In contrast to photosynthetic activity under UV-A radiation, the induction of the biosynthesis of M-Gly was significantly more prominent under UV-B radiation. The content of M-Gly was found to increase with the increase in exposure time under UV-B radiation. The MAA M-Gly was highly stable under UV radiation, heat, strongly acidic and alkaline conditions. It also exhibited good antioxidant activity and photoprotective ability by detoxifying the in vivo reactive oxygen species (ROS) generated by UV radiation. Our results indicate that the studied cyanobacterium may protect itself by synthesizing the UV-absorbing/screening compounds as important defense mechanisms, in their natural brightly-lit habitat with high solar UV-B fluxes.

Temporal dynamics of ROS biogenesis under simulated solar radiation in the cyanobacterium Anabaena variabilis PCC 7937.

We studied the temporal generation of reactive oxygen species (ROS) in the cyanobacterium Anabaena variabilis PCC 7937 under simulated solar radiation using WG 280, WG 295, WG 305, WG 320, WG 335, WG 345, and GG 400 nm cut-off filters to find out the minimum exposure time and most effective region of the solar spectrum inducing highest level of ROS. There was no significant generation of ROS in all treatments in comparison to the samples kept in the dark during the first 8 h of exposure; however, after 12 h of exposure, ROS were significantly generated in samples covered with 305, 295, or 280 nm cut-off filters. In contrast with ROS, the fragmentation of filaments was predominantly seen in 280 nm cut-off filter covered samples after 12 h of exposure. After 24 h of exposure, ROS levels were significantly higher in all samples than in the dark; however, the ROS signals were more pronounced in 320, 305, 295, or 280 nm cut-off filter covered samples. In contrast, the length of filaments was reduced in 305, 295, or 280 nm cut-off filter covered samples after 24 h of exposure. Thus, fragmentation of the filament was induced by all wavelengths of the UV-B region contrary to the UV-A region where only shorter wavelengths were able to induce the fragmentation. In contrast, ROS were generated by all wavelengths of the solar spectrum after 24 h of exposure; however, shorter wavelengths of both the UV-A and the UV-B regions were more effective in generating ROS in comparison to their higher wavelengths and photosynthetic active radiation (PAR). Moreover, lower wavelengths of UV-B were more efficient than the lower wavelengths of the UV-A radiation. Findings from this study suggest that certain threshold levels of ROS are required to induce the fragmentation of filaments.

Characterization of UV-screening compounds, mycosporine-like amino acids, and scytonemin in the cyanobacterium Lyngbya sp. CU2555.

Ultraviolet-screening compounds from the cyanobacterium Lyngbya sp. CU2555 were partially characterized and investigated for their induction by UV radiation, stability under different abiotic factors, and free radical scavenging activity. Based on the high-performance liquid chromatography coupled with diode array detector and ion trap liquid chromatography/mass spectrometry analysis, the compounds were identified as palythine (UVλ max: 319 nm; m/z: 245), asterina (UVλ max: 330 nm; m/z: 289), scytonemin (UVλ max: 384 nm; mw: 544), and reduced scytonemin (UVλ max: 384 nm; m/z: 547). This is the first report for the occurrence of palythine, asterina, and an unknown mycosporine-like amino acids (MAA), M-312 (UVλ max: 312 ± 1 nm), in addition to scytonemin and reduced scytonemin in Lyngbya strains studied so far. Induction of MAAs and scytonemin was significantly more prominent upon exposure to UV-A + UV-B radiation. Both MAAs and scytonemin were highly resistant to some physicochemical factors such as UV-B, heat, and a strong oxidizing agent and exhibited strong antioxidant activity. These results indicate that the studied cyanobacterium may protect itself from deleterious short-wavelength radiation by synthesizing photoprotective compounds in response to harmful UV radiation.

UV radiation-induced biosynthesis, stability and antioxidant activity of mycosporine-like amino acids (MAAs) in a unicellular cyanobacterium Gloeocapsa sp. CU2556.

The biosynthesis of natural sunscreening compounds as influenced by ultraviolet radiation, their stability and antioxidant activity were studied in the cyanobacterium Gloeocapsa sp. CU-2556. An analysis by high-performance liquid chromatography (HPLC) with photodiode-array (PDA) detection revealed the biosynthesis of two MAAs, shinorine (UVλmax 333nm) and an unknown MAA designated as M-307 (UVλmax 307nm) with retention times of 5.9 and 6.4min, respectively. Induction of the synthesis of MAAs was studied under 395 (PAR), 320 (PAR+UV-A) and 295 (PAR+UV-A+UV-B) nm cut-off filters. MAAs induction was significantly increased with an increase in exposure time up to 72h in the samples covered with 295nm cut-off filters. Contrary to shinorine, the biosynthesis of M-307 was more dominant in this unicellular cyanobacterium. Both MAAs were highly stable to some physico-chemical stressors such as UV radiation, heat and a strong oxidizing agent. The MAA M-307 was more stable under strong oxidative stress than shinorine. Moreover, UV-C radiation drastically decreased the stability of both MAAs. The MAAs (shinorine+M-307) also exhibited efficient antioxidant activity which was dose-dependent. The results indicate that MAAs may perform a vital role in survival and sustainability of Gloeocapsa sp. CU-2556 in harsh environmental conditions by its ability to absorb/screen short wavelength UV radiation and antioxidant function.

Partial characterization, UV-induction and photoprotective function of sunscreen pigment, scytonemin from Rivularia sp. HKAR-4.

Scytonemin, located in the extracellular polysaccharide sheath of some cyanobacterial species is considered an efficient natural photoprotectant against lethal doses of ultraviolet (UV) radiations. In the present study, scytonemin from the cyanobacterium Rivularia sp. HKAR-4 was partially characterized and investigated for its induction by UV radiation as well as its role in photoprotection. High-performance liquid chromatography (HPLC) with photodiode-array detection studies revealed the presence of an UV-absorbing compound with absorption maximum at 386 nm. Based on its absorption spectrum and ion trap liquid chromatography/mass spectrometry (LC/MS) analysis, the compound was confirmed as scytonemin. In comparison to photosynthetically active radiation, a significant induction in the synthesis of scytonemin was found under UV-stress. Scytonemin also exhibited efficient photoprotective ability by detoxifying the in vivo reactive oxygen species (ROS) generated by UV radiation and by reducing the formation of thymine dimers. To the best of our knowledge this is the first report on the UV-screening effects of scytonemin on in vivo ROS generation and thymine dimer formation in any cyanobacterial strain. Based on these findings, we conclude that scytonemin may play a vital role in the survival and sustainability of cyanobacterial life in adverse environmental conditions such as under high solar irradiances.

UV radiation-induced accumulation of photoprotective compounds in the green alga Tetraspora sp. CU2551.

The effect of UV radiation on the accumulation of novel mycosporine-like amino acids (MAAs) along with their photoprotective function was investigated in the green alga Tetraspora sp. CU2551. No UV-absorbing compound was detected in this organism growing under normal light condition while two MAAs with absorption maxima at 324 nm and 322 nm were found to be accumulated after UV irradiation. The effects of UV exposure time with different cut-off filter foils namely 295 (PAR + UV-A + UV-B), 320 (PAR + UV-A) and 395 nm (PAR only) were studied on induction of the synthesis of these MAAs. Concentration of MAAs was found to increase with increase in exposure time under UV radiation. Furthermore, the antioxidant and photoprotective action of these MAAs was also investigated. The role of MAAs in diminishing the UV-induced production of ROS in vivo was also demonstrated using the oxidant-sensing probe 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) and results obtained supported the results of DPPH free radical scavenging assay. The MAAs also exhibited efficient photoprotective ability on Escherichia coli cells against UV-B stress. Thus, the MAAs in Tetraspora sp. CU2551 may act as efficient antioxidants as well as UV-sunscreen. This is the first report for the UV-induced synthesis and co-accumulation of these MAAs and their photoprotective actions in Tetraspora sp. which is a member of the class Chlorophyceae. Moreover, UV-induced accumulation as well as photoprotective function of these compounds may facilitate this chlorophyte to perform important ecological functions in harsh environmental conditions with high UV-B fluxes in their brightly lit habitats.

Molecular characterization of hot spring cyanobacteria and evaluation of their photoprotective compounds.

Phylogenetic analysis of 4 cyanobacterial strains isolated from hot springs in Rajgir, India, was carried out using the 16S rRNA gene (1400 bp). These strains were identified as members of Chroococcales ( Cyanothece sp. strain HKAR-1) and Nostocales ( Nostoc sp. strain HKAR-2, Scytonema sp. strain HKAR-3, and Rivularia sp. strain HKAR-4). Furthermore, we evaluated the presence of ultraviolet-screening and (or) photoprotective compounds, such as mycosporine-like amino acids (MAAs) and scytonemin, in these cyanobacteria by using high-performance liquid chromatography. Well-characterized MAAs, including the critical and highly polar compounds shinorine, porphyra-334, and mycosporine-glycine, as well as several unknown MAAs, were found in these hot-spring-inhabiting microorganisms. The presence of scytonemin was detected only in Scytonema sp. strain HKAR-3 and Rivularia sp. strain HKAR-4. The results indicate that hot spring cyanobacteria, namely Cyanothece, Nostoc, Scytonema, and Rivularia, belonging to different groups possess various photoprotective compounds to cope up with the negative impacts of damaging radiations.

Detection of reactive oxygen species (ROS) by the oxidant-sensing probe 2',7'-dichlorodihydrofluorescein diacetate in the cyanobacterium Anabaena variabilis PCC 7937.

The generation of reactive oxygen species (ROS) under simulated solar radiation (UV-B: 0.30Wm(-2), UV-A: 25.70Wm(-2) and PAR: 118.06Wm(-2)) was studied in the cyanobacterium Anabaena variabilis PCC 7937 using the oxidant-sensing fluorescent probe 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA). DCFH-DA is a nonpolar dye, converted into the polar derivative DCFH by cellular esterases that are nonfluorescent but switched to highly fluorescent DCF when oxidized by intracellular ROS and other peroxides. The images obtained from the fluorescence microscope after 12h of irradiation showed green fluorescence from cells covered with 295, 320 or 395nm cut-off filters, indicating the generation of ROS in all treatments. However, the green/red fluorescence ratio obtained from fluorescence microscopic analysis showed the highest generation of ROS after UV-B radiation in comparison to PAR or UV-A radiation. Production of ROS was also measured by a spectrofluorophotometer and results obtained supported the results of fluorescence microscopy. Low levels of ROS were detected at the start (0h) of the experiment showing that they are generated even during normal metabolism. This study also showed that UV-B radiation causes the fragmentation of the cyanobacterial filaments which could be due to the observed oxidative stress. This is the first report for the detection of intracellular ROS in a cyanobacterium by fluorescence microscopy using DCFH-DA and thereby suggesting the applicability of this method in the study of in vivo generation of ROS.