Regulation of redox homeostasis in cadmium stressed rice field cyanobacteria by exogenous hydrogen peroxide and nitric oxide.

In the present study, defensive strategies of H2O2 mediated NO signaling were analyzed in Cd stressed Nostoc muscorum and Anabaena sp. Exogenously supplied SNP (10 µM) and H2O2 (1 µM) lessen the toxicity of Cd (6 µM) but without NO; H2O2 was unable to release the stress from cyanobacterial cells potentially. The reduced contents of exopolysaccharide, protein content, endogenous NO and enzymatic antioxidants (SOD, POD, CAT, and GST) due to Cd toxicity, were found increased significantly after exogenous application of H2O2 and SNP thereafter, cyanobacterial calls flourished much better after releasing toxic level of Cd. Moreover, increased level of ROS due to Cd stress also normalized under exogenous application of H2O2 and SNP. However, chelation of NO hindered the signaling mechanism of H2O2 that diminished its potential against Cd stress while signaling of NO has not been hindered by chelation of H2O2 and NO potentially released the Cd stress from cyanobacterial cells. In conclusion, current findings demonstrated the synergistic signaling between H2O2 and NO towards the improvement of cyanobacterial tolerance to Cd stress, thereby enhancing the growth and antioxidant defense system of test cyanobacteria that improved fertility and productivity of soil even under the situation of metal contamination.

Analyzing dose dependency of antioxidant defense system in the cyanobacterium Nostoc muscorum Meg 1 chronically exposed to Cd2.

The aim of the present study was to analyze the dose dependency of oxidant-antioxidant homeostasis in Cd2+ exposed Nostoc muscorum Meg 1 cells. Quantification of percent DNA loss, protein oxidation and lipid peroxidation was carried out to assess Cd2+ induced ROS mediated damages to the organism. The countermeasures adopted by the cyanobacterium were also evaluated by computing various components of both enzymatic and non-enzymatic antioxidants. Exposure to different Cd2+ (0.1, 0.2, 0.3, 0.5, 1, 1.5, 2, 2.5, 3 ppm) doses showed substantial increase in ROS content in the ranges of 20-181% and 116-323% at the end of first and seventh day. The DNA damage, protein oxidation and lipid peroxidation were increased by 11-62%, 7-143% and 13-183% with increasing Cd2+ concentrations at the end of seven days. TEM images clearly showed damages to the cell wall, cell membrane and thylakoid organization at higher Cd2+ (0.5-3 ppm) concentrations. Cd2+ exposure up to 0.5 ppm registered increase in contents of antioxidative enzymes (superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione reductase (GR)) and in non-enzymatic antioxidants (glutathione, total thiol, phytochelatin and proline) indicating stimulation of ROS mitigating machinery. However, toxicity of Cd2+ was evident as at higher concentrations the cellular morphology and ultra-structures were negatively affected and the capacities of the cells to generate various antioxidant measures were highly compromised. The organism registered 96-98% sorption ability from a solution supplemented with 0.3 ppm Cd2+ and thus show realistic potential as Cd2+ bioremediator in wastewater treatment.

Cytokinin alleviates cypermethrin toxicity in Nostoc muscorum by involving nitric oxide: Regulation of exopolysaccharides secretion, PS II photochemistry and reactive oxygen species homeostasis.

Growth of the most important nitrogen fixing cyanobacterium Nostoc muscorum is reported to be badly affected by the application of insecticides. To overcome their damaging effects, several strategies are being used. Out of these, some works on kinetin (KN, a synthetic cytokinin) has been recognized that it can overcome toxicity of insecticides in cyanobacteria. Besides this, it is now known that every hormone needs certain second messengers such as nitric oxide (NO) for its action. But implication of NO in KN-mediated regulation of insecticide toxicity is yet to be investigated. Hence in the current study, we have investigated the possible involvement of NO in KN-mediated regulation of cypermethrin toxicity in the cyanobacterium Nostoc muscorum. Cypermethrin decreased growth of Nostoc muscorum which was accompanied by decreased pigment contents and altered photosystem II (PS II) photochemistry that resulted in inhibition of photosynthetic process but KN significantly ameliorated cypermethrin toxicity. Cypermethrin induced production of free radicals (in-vivo and in-vitro) and weakened defensive mechanism (enzymatic and non-enzymatic defense system) which was restored by KN. Further, the results revealed that NG-nitro-l-arginine methyl ester (l-NAME, an inhibitor of nitric oxide synthase) worsened the effect of cypermethrin toxicity even in the presence of KN while 2-4-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO, a scavenger of NO) reversed KN-mediated amelioration even in the presence of sodium nitroprusside (SNP, an NO donor), suggesting that endogenous NO is required for mitigation of cypermethrin toxicity. Overall, our results first time show that endogenous NO is essential for KN-mediated mitigation of cypermethrin toxicity in the Nostoc muscorum.

Comparative studies on growth and Pb(II) removal from aqueous solution by Nostoc muscorum and Anabaena variabilis.

Industrial wastewater containing heavy metals is a major environmental problem that needs to be treated. This study reported the ability of two fresh water algae cyanobacteria (Nostoc muscorum and Anabaena variabilis) to remove lead from aqueous solutions of four different initial concentrations (0-50 mg/L-1) for 21 days under controlled laboratory conditions. Results obtained in this study showed a maximum removal of Pb(II) (97.8%) by N. muscorum at 15 mg/L-1 initial metal concentration however the maximum removal by A. variabilis at the same concentration was 71.4% after 16 day of incubation. These N. muscorum appeared to be more efficient than A. variabilis for removing Pb(II). Algal growth, pigments in the algae cells were measured during incubation period. Lower concentrations of lead increased biomass, OD, chlorophyll a and carotenoids in both algae. On the other hand, higher concentrations of lead were inhibitory for growth.

Kinetin alleviates chromium toxicity on growth and PS II photochemistry in Nostoc muscorum by regulating antioxidant system.

The present study was undertaken to evaluate the metal toxicity alleviating effects of kinetin (KN, 10 nM) on growth, photosynthetic pigments and photochemistry of PS II in the cyanobacterium Nostoc muscorum exposed to chromium (CrVI) stress (100 and 150 µM). Chromium declined growth, photosynthetic pigments (chlorophyll a, phycocyanin and carotenoids), photosynthetic oxygen evolution rate and parameters of fluorescence kinetics (ϕP0, FV/F0, ϕE0, Ψ0 and PIABS except F0/FV) in concentration dependent manner, while stimulating effects on respiration, energy flux parameters (ABS/RC, TR0/RC, ET0/RC and DI0/RC), oxidative stress biomarkers i.e., superoxide radical (SOR), hydrogen peroxide (H2O2) and lipid peroxidation (TBARS contents) and antioxidative enzymes: superoxide dismutase (SOD), peroxidase (POD), catalase (CAT) and glutathione-S-transferase (GST), were observed. However, upon addition of KN in the growth medium an alleviating effect against chromium induced toxicity on growth, photosynthetic pigments and photochemistry of PS II was recorded. This had occurred due to substantial reduction in levels of oxidative stress biomarkers: SOR, H2O2 and TBARS contents with concomitant rise in activity of antioxidative enzymes: SOD, POD, CAT and GST and appreciable lowering in the cellular accumulation of chromium. The overall results demonstrate that KN application significantly alleviated chromium induced toxicity on growth performance of the cyanobacterium N. muscorum due to significant improvement in photosynthetic pigments and photochemistry of PS II by up-regulating the activity of antioxidative enzymes, and declining cellular accumulation of chromium. Furthermore, Cr induced toxicity at lower dose (100 µM) was found to be ameliorated more efficiently in N. muscorum following supplementation of KN.

Toxicity assessment of arsenate and arsenite on growth, chlorophyll a fluorescence and antioxidant machinery in Nostoc muscorum.

The present study deals with impact of varied doses of arsenite (AsIII; 50, 100 and 150 µM) and arsenate (AsV; 50, 100 and 150 mM) on growth, photosynthetic pigments, photochemistry of photosystem II, oxidative biomarkers, (O2•¯, H2O2 and MDA equivalents contents) and activity of antioxidant enzymes in diazotrophic cyanobacterium Nostoc muscorum after 48 and 96 h of the treatments. The reduction in growth, pigment contents (Chl a, Phy and Car) and PS II photochemistry was found to increase with enhanced accumulation of test metal in cells, and the damaging effect on photosynthetic pigments showed the order (Phy > chl a> Car). The negative effect on PS II photochemistry was due to significant decrease in the value of JIP kinetics ϕP0, FV/F0, ϕE0,Ψ0 and PIABS except F0/FV and significant rise in values of energy flux parameters such as ABS/RC, TR0/RC, ET0/RC and DI0/RC. Both the species of arsenic caused significant rise in oxidative biomarkers as evident by in vitro and in vivo analysis of (O2•¯, H2O2 and MDA equivalents contents) despite of appreciable rise in the activity antioxidative enzymes such as SOD, POD, CAT and GST. The study concludes that in among both forms of arsenic, arsenite effect was more dominant on growth, photosynthetic pigments; oxidative stress biomarkers as evident by weak induction of anti-oxidative defense system to overcome the stress as compared to arsenate.

Pretilachlor toxicity is decided by discrete photo-acclimatizing conditions: Physiological and biochemical evidence from Anabaena sp. and Nostoc muscorum.

The current study was undertaken to elucidate the impact of the herbicide pretilachlor (3 µg ml-1 and 6 µg ml-1) on cyanobacteria, Nostoc muscorum ATCC 27893 and Anabaena sp. PCC 7120 under three levels of photoacclimatization (suboptimum, 25 µmol photon m-2 s-1; optimum, 75 µmol photon m-2 s-1; and supra-optimum, 225 µmol photon m-2 s-1) by analyzing certain physiological (biomass accumulation, photosynthesis, Chl a fluorescence and respiration) and biochemical parameters (photosynthetic pigments‒ chlorophyll a, carotenoids and phycocyanin; reactive oxygen species‒ O2•¯, H2O2, lipid peroxidation; antioxidant system‒ superoxide dismutase, peroxidise, catalase and glutathione-S-transferase). The light conditioning played the most prominent role in deciding the extent of herbicide toxicity on both the tested cyanobacteria as the maximum toxicity was observed in suboptimum light acclimatized cyanobacterial cells corroborated by the least growth in the same cells. The impact of pretilachlor treatment on photosystem II photochemistry viz. φP0, Ñ°0, φE0, PIABS, ABS/RC, TR0/RC, ET0/RC and DI0/RC was also altered by light acclimatization. The percent rise in oxidative stress markers (SOR and H2O2) and consequent lipid peroxidation (MDA equivalents) were also highest in suboptimum light acclimatized cells exposed to pretilachlor which could not be prospered with compatible antioxidant performance. Conversely, supra-optimum light acclimatized cells of both the cyanobacteria was found to accelerate the activities of all the studied enzymes and thus able to counterbalance the pretilachlor toxicity and supported the healthier growth.

NaCl-induced physiological and biochemical changes in two cyanobacteria Nostoc muscorum and Phormidium foveolarum acclimatized to different photosynthetically active radiation.

The present study is aimed at investigating physiological and biochemical behavior of two cyanobacteria Nostoc muscorum and Phormidium foveolarum acclimatized to different levels (sub-optimum; 25 ± 0.5, optimum; 75 ± 2.5 and supra-optimum; 225 ± 3.5 µmol photons m(-2) s(-1)) of photosynthetic active radiation (PAR), and subsequently treated with two doses (30 and 90 mM) of NaCl. PAR influences growth in tested cyanobacteria being maximum in supra-optimum PAR acclimatized cells. NaCl-induced maximum percent decline in growth was observed in sub-optimum PAR acclimatized cells, which was in consonance with a decrease in chlorophyll content. Sub-optimum PAR acclimatization stimulated phycocyanin content in control cells, whereas maximum carotenoids content was observed in supra-optimum PAR acclimatized cells. Photosystem II photochemistry viz. Fv/F0, Fv/Fm, Ψ0, ϕE0, PIABS, ABS/RC, TR0/RC, ET0/RC and DI0/RC was also influenced by PAR and NaCl. Maximum percent rise in superoxide radical (SOR), hydrogen peroxide (H2O2) and lipid peroxidation was observed in sub-optimum PAR acclimatized cells exposed to NaCl, which could be correlated with lower values of enzymatic (superoxide dismutase, catalase, peroxidase and glutathione-S-transferase) and non-enzymatic (NP-SH and cysteine) antioxidants. In supra-optimum PAR acclimatized cells level of oxidative stress markers was in parallel with enhanced antioxidants. The results suggest that PAR significantly changes physiological and biochemical responses of studied cyanobacteria under NaCl stress. Besides this, this study also shows that P. foveolarum is more tolerant than N. muscorum under test conditions.

Low and high doses of UV-B differentially modulate chlorpyrifos-induced alterations in nitrogen metabolism of cyanobacteria.

The present study assessed the comparative responses on the specific growth rate, nitrogen metabolism and enzymes associated with nitrogen metabolism in two nitrogen fixing cyanobacteria-Nostoc muscorum and Phormidium foveolarum exposed to two UV-B doses (low; UV-BL: 0.5472kJm(-2) and high; UV-BH: 5.472kJm(-2)) and two doses of the insecticide chlorpyrifos (O,O-diethyl O-3,5,6-trichloro-2-pyridyl phosphorothioate; low i.e. CPL, 1µgml(-1) and high i.e. CPH, 2µgml(-1)) singly and in combination. The specific growth rate, NO3(-) and NO2(-) uptake, nitrate assimilating enzymes - nitrate reductase and nitrite reductase and ammonium assimilating enzymes - glutamine synthetase and glutamate synthase were severely affected when treated either with CPH or/and UV-BH while glutamate dehydrogenase exhibited a stimulatory response. CPL also reduced all the measured parameters (except GDH activity) after 24h, however, a stimulatory effect was observed after 72h due to an increase in nitrogen metabolism (and other antioxidant) enzymes during this period. UV-BL did not cause significant alteration in the studied parameters while in combination with CP doses, it either alleviated the inhibitory effects or further enhanced the CPL induced activities of these enzymes (except GDH). Overall results indicate the resistant nature of P. foveolarum against the inhibitory doses of UV-B and chlorpyrifos in comparison to N. muscorum.

Compatibility of ascorbate-glutathione cycle enzymes in cyanobacteria against low and high UV-B exposures, simultaneously exposed to low and high doses of chlorpyrifos.

This study deals with the comparative responses of the two cyanobacteria viz. Nostoc muscorum and Phormidium foveolarum against single and combined doses of low (UV-B(L,) 0.1 µmol m(-2) s(-1)) and high (UV-B(H), 1.0 µmol m(-2) s(-1)) fluence rates of ultraviolet-B radiation with low (CP(L), 1 µg ml(-1)) and high (CP(H), 2 µg ml(-1)) doses of the insecticide chlorpyrifos by measuring changes in growth, ascorbate-glutathione cycle enzymes and related metabolites. CP(L) and UV-B(L) both caused lesser increase in ROS but significantly stimulated AsA-GSH cycle enzymes. On the other hand, CP(H) and UV-B(H) posed inhibitory effects by enhancing ROS and inhibiting AsA-GSH cycle enzymes. Inhibitions in CP(H) or UV-B(H) treated samples were significantly prevented when they were supplemented with UV-B(L) and CP(L) (after 72 h), respectively by lowering down ROS and enhancing AsA-GSH enzymes and related metabolites which manifested in terms of improved biomass accumulation.

High light intensity augments mercury toxicity in cyanobacterium Nostoc muscorum.

The present study is aimed at investigating the role of growth irradiance in determining the extent of mercury (Hg) toxicity on various physiological parameters viz. growth, pigment contents, photosynthesis, respiration, (14)CO(2) fixation, photosynthetic electron transport, photorespiration and enzyme activity of cyanobacterium Nostoc muscorum. A general decline was observed in all these parameters with increasing concentration of Hg except for carotenoids content and respiratory activity which exhibited significant enhancement. This effect was more pronounced in high light (130 µmol photon m(-2) s(-1)) exposed cells as compared to normal (70 µmol photon m(-2) s(-1)) and low (10 µmol photon m(-2) s(-1)) light exposed cells. Among the photosynthetic electron transport activities, whole chain was found to be more sensitive than photosystem II (PSII) and photosystem I (PSI). (14)CO(2) fixation was more affected as compared to O(2) evolution when exposed to Hg and different light intensities. Photorespiratory activity, which is an index of protecting organisms from light-induced damage, also showed a similar declining trend. Enzyme assay revealed that among the carboxylating enzymes, activity of RUBISCO was more severely inhibited than PEPCase. Thus, these results suggest that Hg itself was toxic at all tested concentrations and high light intensity augmented its toxicity in N. muscorum inhibiting the growth, pigment contents and photosynthetic activity of the organism.

UV-B induced differential effect on growth and nitrogen metabolism in two cyanobacteria under copper toxicity.

In the present study, impact of low (UV-B(L): 0.1 µmol m(-2) s(-1)) and high (UV-BH: 1.0 µmol m(-2) s(-1)) fluence rates of ultraviolet-B on growth and nitrogen metabolism in two cyanobacteria: Phormidium foveolarum and Nostoc muscorum under copper toxicity (2 and 5 µM) was investigated after 24 and 72 h of experiments. Copper and UV-BH treatment suppressed growth but more in N. muscorum which was accompanied by significant accumulation of Cu. Nitrate and nitrite uptake rates and activities of nitrogen assimilating enzymes i.e. nitrate reductase (NR), nitrite reductase (NiR), glutamine synthetase (GS) and glutamate synthase (GOGAT) except glutamate dehydrogenase activity (GDH; aminating) were decreased following treatments of Cu and UV-BH, and under combined treatments the effect was greater. On contrary, UV-BL declined Cu toxicity significantly. The study concludes that Cu and UV-BH suppressed the activity of NR, NiR, GS and GOGAT (except GDH) hence decreased growth. However, UV-BL showed cross tolerance in test organisms against Cu toxicity up to certain extent. Phormidium foveolarum is comparatively less sensitive against UV-BH and excess Cu, a situation likely exists in nature, hence it may be used as a biofertilizer for sustainable agriculture.

Differential physiological and biochemical responses of two cyanobacteria Nostoc muscorum and Phormidium foveolarum against oxyfluorfen and UV-B radiation.

In the present study, degree of tolerance and tolerance strategies of two paddy field cyanobacteria viz. Nostoc muscorum and Phormidium foveolarum against oxyfluorfen (10 and 20 µg ml(-1)) and UV-B (7.2 kJ m(-2)d(-1)) stress were investigated. Oxyfluorfen and UV-B decreased growth, photosynthesis, nutrient uptake, nitrate reductase, acid and alkaline phosphatase activities, which accompanied with the increase in the level of oxidative stress. However, growth was more affected in N. muscorum than P. foveolarum. Antioxidants exhibited differential responses against oxyfluorfen and UV-B stress. Ascorbate and proline levels were higher in P. foveolarum. A protein of 66 kDa was expressed in N. muscorum, however, it was absent in P. foveolarum than those of N. muscorum. Besides this, a protein of 29 kDa appeared in P. foveolarum under all the treatments, but it was present only in control cells of N. muscorum cells. Overall results indicated resistant nature of P. foveolarum against oxyfluorfen and UV-B stress in comparison to N. muscorum.

Comparative toxicity of bentazon and molinate on growth, photosynthetic pigments, photosynthesis, and respiration of the Portuguese ricefield cyanobacterium Nostoc muscorum.

Bentazon and molinate are selective herbicides recommended for integrated weed management in rice. Their toxicity on growth and some biochemical and physiological parameters of Nostoc muscorum, an abundant cyanobacterium in Portuguese rice fields, was evaluated under laboratory conditions during time- and concentration-dependent exposure for 72 h. Results showed that toxic concentrations (0.75-2 mM) of both herbicides have pleiotropic effects on the cyanobacterium. Molinate was more toxic than bentazon to growth, respiration, chlorophyll-a, carotenoids, and phycobiliproteins contents. Protein content was increased by both herbicides although the effect was particularly evident with higher concentrations of molinate (1.5-2 mM). The herbicides had contrasting effects on carbohydrates content: molinate increased this organic fraction whereas bentazon decreased it. Photosynthesis and respiration were inhibited by both herbicides.

Toxicity assessment of heavy metals with Nostoc muscorum L.

Heavy metals in aquatic ecosystem (Mn, Ni, Zn, Cu) were tested for their toxicity against Nostoc muscorum L., a common fresh water phytoplankton, with respect to chlorophyll, protein, total carbohydrate and starch contents along with growth (OD). Mn showed stimulatory effect up to 1.5 ppm for all the observed parameters. However, Ni, Zn and Cu (0.1, 0.2, 0.3, 0.4 and 0.5 ppm) showed adverse effects even at 0.1 ppm. Chlorophyll was most sensitive parameter followed by carbohydrate, protein and starch. Ni was most toxic to N. muscorum followed by Cu and Zn. The IC50 for Ni was 0.1 ppm for growth, chlorophyll and protein, while total carbohydrate and starch showed IC50 at 0.3 and 0.4 ppm, respectively. The IC50 for Cu was recorded at 0.2 ppm for all the parameters. Zn showed IC50 at 0.3 ppm for growth, chlorophyll, protein and starch at 0.4 ppm for carbohydrate. This study can be applied as bioassay using cyanobacteria for toxicity assessment of various industrial wastes in aquatic ecosystem.

Poly-beta-hydroxybutyrate accumulation in Nostoc muscorum and Spirulina platensis under phosphate limitation.

Nostoc muscorum and Spirulina platensis were grown under phosphate deficiency in order to investigate the role of internal phosphate pool and activity of alkaline phosphatase on poly-beta-hydroxybutyrate (PHB) accumulation. PHB accumulation in N. muscorum increased to 22.7% of dry weight (dw) after 4 day of phosphate deficiency, while the internal phosphate pool reduced to the level of 0.02 microM mg dw(-1) at a maximum APase activity of 2.57nM PNP mg dw(-1) h(-1). In contrary, S. platensis depicted maxima of 1.39nM PNP mg dw(-1) h(-1) on day 30 of incubation, which was about 2 fold lower than the observed value of N. muscorum. PHB content in S. platensis remained low even after prolonged phosphate starvation, and a rise only up to 3.5% of dw was recorded on day 60 of phosphate deficiency. Supplementation of NADPH exogenously to S. platensis cultures grown under phosphate deficiency favoured PHB accumulation in 10, 20 and 30 days old cultures, but not in the cultures grown under phosphate deficiency for 60 days. The possible role of phosphate limitation on PHB accumulation is discussed.