Testing the Potential of Regulatory Sigma Factor Mutants for Wastewater Purification or Bioreactor Run in High Light.

It is shown that a freshly inoculated culture of the model cyanobacterium Synechocystis sp. PCC 6803 consumed almost all phosphate and 50% of nitrate within 6 days from the nutrient-rich BG-11 growth medium, indicating potential of cyanobacteria to purify wastewaters. Synechocystis sp. PCC 6803 control strain also collected nutrients efficiently from a landfill leachate wastewater KA2 (5.9-6.9 mM ammonium and 0.073-0.077 mM phosphate). Wastewaters might induce oxidative stress to microalgae, which prompted us to test growth of sigma factor inactivation strains, as ΔsigBCE and ΔsigCDE strains show superior growth in chemically induced oxidative stress. All cyanobacterial strains, including a stress-sensitive strain ΔsigBCDE, grew well in KA2 for four days, indicating that KA2 did not cause immediate oxidative stress. Completely arrested growth and bleaching of ΔsigBCDE cells after one week in KA2 wastewater point to the importance of group 2 sigma factor-mediated changes in gene expression during wastewater treatment. The growth of ΔsigBCD was arrested early in un-buffered and Hepes buffered (pH 7.5) KA2. In ΔsigBCD, all phosphate transporter genes are upregulated in standard conditions, and ΔsigBCD cells showed growth defects in low-phosphate BG-11 medium. ΔsigBCD cells removed phosphate slower from KA2 than the control strain, but phosphate supplementation of KA2 did not improve growth of ΔsigBCD. The ΔsigBCE strain showed superior growth in a laboratory-scale bioreactor in bright light and removed phosphate even slightly more efficiently than the control strain if KA2 was Hepes buffered although ΔsigBCE grew slowly in un-buffered KA2 and in low-phosphate BG-11 medium. The results indicate that engineering expression of regulatory group 2 sigma factor(s) might be useful for practical applications.

Group 2 Sigma Factors are Central Regulators of Oxidative Stress Acclimation in Cyanobacteria.

Regulatory σ factors of the RNA polymerase (RNAP) adjust gene expression according to environmental cues when the cyanobacterium Synechocystis sp. PCC 6803 acclimates to suboptimal conditions. Here we show central roles of the non-essential group 2 σ factors in oxidative stress responses. Cells missing all group 2 σ factors fail to acclimate to chemically induced singlet oxygen, superoxide or H2O2 stresses, and lose pigments in high light. SigB and SigD are the major σ factors in oxidative stress, whereas SigC and SigE play only minor roles. The SigD factor is up-regulated in high light, singlet oxygen and H2O2 stresses, and overproduction of the SigD factor in the ΔsigBCE strain leads to superior growth of ΔsigBCE cells in those stress conditions. Superoxide does not induce the production of the SigD factor but instead SigB and SigC factors are moderately induced. The SigB factor alone in ΔsigCDE can support almost as fast growth in superoxide stress as the full complement of σ factors in the control strain, but an overdose of the stationary phase-related SigC factor causes growth arrest of ΔsigBDE in superoxide stress. A drastic decrease of the functional RNAP limits the transcription capacity of the cells in H2O2 stress, which explains why cyanobacteria are sensitive to H2O2. Formation of RNAP-SigB and RNAP-SigD holoenzymes is highly enhanced in H2O2 stress, and cells containing only SigB (ΔsigCDE) or SigD (ΔsigBCE) show superior growth in H2O2 stress.

Degradation of chlorophyll and synthesis of flavonols during autumn senescence-the story told by individual leaves.

Autumn senescence of deciduous trees is characterized by chlorophyll degradation and flavonoid synthesis. In the present study, chlorophyll and flavonol contents were measured every morning and evening during the whole autumn with a non-destructive method from individual leaves of Sorbus aucuparia, Acer platanoides, Betula pendula and Prunus padus. In most of the studied trees, the chlorophyll content of each individual leaf remained constant until a phase of rapid degradation commenced. The fast phase lasted only ~1 week and ended with abscission. In S. aucuparia, contrary to the other species, the chlorophyll content of leaflets slowly but steadily decreased during the whole autumn, but rapid chlorophyll degradation commenced only prior to leaflet abscission also in this species. An increase in flavonols commonly accompanied the rapid degradation of chlorophyll. The results may suggest that each individual tree leaf retains its photosynthetic activity, reflected by a high chlorophyll content, until a rapid phase of chlorophyll degradation and flavonoid synthesis begins. Therefore, in studies of autumn senescence, leaves whose chlorophyll content is decreasing and leaves with summertime chlorophyll content (i.e. the leaves that have not yet started to degrade chlorophyll) should be treated separately.

Cultivation of Nannochloropsis for eicosapentaenoic acid production in wastewaters of pulp and paper industry.

The eicosapentaenoic acid (EPA) containing marine microalga Nannochloropsis oculata was grown in an effluent from anaerobic digestion of excess activated sludge from a wastewater treatment plant serving a combination of a pulp and a paper mill and a municipality (digester effluent, DE), mixed with the effluent of the same wastewater treatment plant. The maximum specific growth rate and photosynthesis of N. oculata were similar in the DE medium and in artificial sea water medium (ASW) but after 7 days, algae grown in the DE medium contained seven times more triacylglycerols (TAGs) per cell than cells grown in ASW, indicating mild stress in the DE medium. However, the volumetric rate of EPA production was similar in the ASW and DE media. The results suggest that N. oculata could be used to produce EPA, utilizing the nutrients available after anaerobic digestion of excess activated sludge of a pulp and paper mill.

Effect of dynamic oxygen concentrations on the growth of Neochloris oleoabundans at sub-saturating light conditions.

In tubular photobioreactors micro-algae continuously experience dynamically changing oxygen and light conditions when circulating from the solar receiver to the dark degasser. These changes in oxygen concentration and light were simulated in a CSTR using sub-saturating light intensity. Elongation of the residence time in the solar receiver from 30 to 300 min was also investigated. Specific growth rates measured at constant low oxygen concentration PO2=0.21 bar were; 1.14 ± 0.06 day(-1) using continuous light 0.80 ± 0.16 day(-1) with 30 min light and 1.09 ± 0.05 day(-1) with 300 min light. The effect of dynamically changing oxygen concentrations from PO2=0.21 to 0.63 bar followed by degassing resulted in similar specific growth rates. The exposure of the algae cells to dark periods in the degasser has a bigger negative impact than the temporary exposure to accumulating oxygen concentrations in the solar receiver. This shows that considerable energy savings for degassing are possible.