Cyanobacteria are prokaryotic organisms characterised by their complex structures and a wide range of pigments. With their ability to fix CO2, cyanobacteria are interesting for white biotechnology as cell factories to produce various high-value metabolites such as polyhydroxyalkanoates, pigments, or proteins. White biotechnology is the industrial production and processing of chemicals, materials, and energy using microorganisms. It is known that exposing cyanobacteria to low levels of stressors can induce the production of secondary metabolites. Understanding of this phenomenon, known as hormesis, can involve the strategic application of controlled stressors to enhance the production of specific metabolites. Consequently, precise measurement of cyanobacterial viability becomes crucial for process control. However, there is no established reliable and quick viability assay protocol for cyanobacteria since the task is challenging due to strong interferences of autofluorescence signals of intercellular pigments and fluorescent viability probes when flow cytometry is used. We performed the screening of selected fluorescent viability probes used frequently in bacteria viability assays. The results of our investigation demonstrated the efficacy and reliability of three widely utilised types of viability probes for the assessment of the viability of Synechocystis strains. The developed technique can be possibly utilised for the evaluation of the importance of polyhydroxyalkanoates for cyanobacterial cultures with respect to selected stressor-repeated freezing and thawing. The results indicated that the presence of polyhydroxyalkanoate granules in cyanobacterial cells could hypothetically contribute to the survival of repeated freezing and thawing.
- Klíčová slova
- Biotechnology, Cyanobacteria, Flow cytometry, Fluorescent viability probes, Stress resistance, Viability assessment,
- MeSH
- fluorescenční barviva * MeSH
- fyziologický stres * MeSH
- mikrobiální viabilita * MeSH
- polyhydroxyalkanoáty metabolismus MeSH
- průtoková cytometrie * metody MeSH
- sinice * fyziologie MeSH
- Synechocystis * fyziologie MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- fluorescenční barviva * MeSH
- polyhydroxyalkanoáty MeSH
Poly(3-hydroxybutyrate) (PHB) is a biobased and biodegradable polymer with properties comparable to polypropylene and therefore has the potential to replace conventional plastics. PHB is intracellularly accumulated by prokaryotic organisms. For the cells PHB functions manly as carbon and energy source, but all possible functions of PHB are still not known. Synechocystis (cyanobacteria) accumulates PHB using light as energy and CO2 as carbon source. The main trigger for PHB accumulation in cyanobacteria is nitrogen and phosphorous depletion with simultaneous surplus of carbon and energy. For the above reasons, obtaining knowledge about external factors influencing PHB accumulation is of highest interest. This study compares the effect of continuous light exposure and day/night (16/8 h) cycles on selected physiology parameters of three Synechocystis strains. We show that continuous illumination at moderate light intensities leads to an increased PHB accumulation in Synechocystis salina CCALA 192 (max. 14.2% CDW - cell dry weight) compared to day/night cycles (3.7% CDW). In addition to PHB content, glycogen and cell size increased, while cell density and cell viability decreased. The results offer new approaches for further studies to gain deeper insights into the role of PHB in cyanobacteria to obtain bioplastics in a more sustainable and environmentally friendly way.
- Klíčová slova
- PHB, Synechocystis, cell size, continuous illumination, day/night cycle, glycogen,
- Publikační typ
- časopisecké články MeSH
Sample preparation protocols for conventional high voltage transmission electron microscopy (TEM) heavily rely on the usage of staining agents containing various heavy metals, most commonly uranyl acetate and lead citrate. However high toxicity, rising legal regulations, and problematic waste disposal of uranyl acetate have increased calls for the reduction or even complete replacement of this staining agent. One of the strategies for uranyless imaging is the employment of low-voltage transmission electron microscopy. To investigate the influence of different imaging and staining strategies on the final image of cyanobacterial cells, samples stained by uranyl acetate with lead citrate, as well as unstained samples, were observed using TEM and accelerating voltages of 200 kV or 25 kV. Moreover, to examine the possibilities of reducing chromatic aberration, which often causes issues when imaging using electrons of lower energies, samples were also imaged using a scanning transmission electron microscopy at 15 kV accelerating voltages. The results of this study demonstrate that low-voltage electron microscopy offers great potential for uranyless electron microscopy.
- Klíčová slova
- Synechocystis, contrasting agents, low voltage electron microscopy, polyhydroxyalkanoates, transmission electron microscopy, uranyl acetate,
- Publikační typ
- časopisecké články MeSH
Polyhydroxyalkanoates (PHA), polyesters accumulated by numerous prokaryotes in the form of intracellular granules, have been for decades considered being predominantly storage molecules. However, numerous recent discoveries revealed and emphasized their complex biological role for microbial cells. Most of all, it was repeatedly reported and confirmed that the presence of PHA granules in prokaryotic cells enhances stress resistance and robustness of microbes against various environmental stress factors such as high or low temperature, freezing, oxidative, and osmotic pressure. It seems that protective mechanisms of PHA granules are associated with their extraordinary architecture and biophysical properties as well as with the complex and deeply interconnected nature of PHA metabolism. Therefore, this review aims at describing novel and unexpected properties of PHA granules with respect to their contribution to stress tolerance of various prokaryotes including common mesophilic heterotrophic bacteria, but also extremophiles or photo-autotrophic cyanobacteria. KEY POINTS: • PHA granules present in bacterial cells reveal unique properties and functions. • PHA enhances stress robustness of bacterial cells.
- Klíčová slova
- Bacteria, Cupriavidus necator, Cyanobacteria, Extremophiles, PHA granules, Polyhydroxyalkanoates, Stress robustness,
- MeSH
- Bacteria metabolismus MeSH
- Cupriavidus necator metabolismus MeSH
- fyziologický stres * MeSH
- osmotický tlak MeSH
- polyhydroxyalkanoáty metabolismus MeSH
- sinice metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
- Názvy látek
- polyhydroxyalkanoáty MeSH
The current commercial production of polyhydroxyalkanoates (PHA) is based on heterotrophic bacteria, using organic carbon sources from crops. To avoid the competition with food and feed production, cyanobacteria, metabolising PHA from carbon dioxide can be used. This research focuses on the investigation of the thermal and rheological properties of PHA polymers accumulated by Synechocystis salina, which had been cultivated in digestate supernatant and a mineral medium. The dried bacterial cells had a polymer content of 5.5-6.6%. The relevance of the derived PHA polymers for the common melt polymer processing was correlated with their molecular mass distribution as well as with their thermal and rheological properties. The determined thermal and rheological properties showed that PHA polymers accumulated by S. salina on digestate supernatant or mineral medium are comparable with the commercial available poly(3-hydroxybutyrate). However, the results showed that PHA polymers in general require modification before melt processing to increase their stability in the molten state.
- Klíčová slova
- Digestate, Polyhydroxyalkanoates, Synechocystis salina,
- MeSH
- kultivační média chemie metabolismus MeSH
- molekulová hmotnost MeSH
- oxid uhličitý metabolismus MeSH
- polyhydroxyalkanoáty biosyntéza chemie MeSH
- Synechocystis metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- kultivační média MeSH
- oxid uhličitý MeSH
- polyhydroxyalkanoáty MeSH
A European round robin test according to ISO 5725-2 was conceptually prepared, realised, and evaluated. The aim was to determine the inter-laboratory variability of the overall process for the ecotoxicological characterization of construction products in eluates and bioassays. To this end, two construction products BAM-G1 (granulate) and HSR-2 (roof sealing sheet), both made of EPDM polymers (rubber), were selected. The granular construction product was eluted in a one stage batch test, the planar product in the Dynamic Surface Leaching test (DSLT). A total of 17 laboratories from 5 countries participated in the round robin test: Germany (12), Austria (2), Belgium (1), Czech Republic (1) and France (1). A test battery of four standardised ecotoxicity tests with algae, daphnia, luminescent bacteria and zebrafish eggs was used. As toxicity measures, EC50 and LID values were calculated. All tests, except the fish egg test, were basically able to demonstrate toxic effects and the level of toxicity. The reproducibility of test results depended on the test specimens and the test organisms. Generally, the variability of the EC50 or LID values increased with the overall level of toxicity. For the very toxic BAM-G1 eluate a relative high variability of CV = 73%-110% was observed for EC50 in all biotests, while for the less toxic HSR-2 eluate the reproducibility of EC50 varied with sensitivity: it was very good (CV = 9.3%) for the daphnia test with the lowest sensitivity, followed by the algae test (CV = 36.4%). The luminescent bacteria test, being the most sensitive bioassay for HSR-2 Eluate, showed the highest variability (CV = 74.8%). When considering the complex overall process the reproducibility of bioassays with eluates from construction products was acceptable.
- Klíčová slova
- Construction products, Ecotoxicity tests, Eluates, Leaching tests, Round robin test,
- MeSH
- Bacteria účinky léků MeSH
- biotest metody normy MeSH
- chemické látky znečišťující vodu analýza toxicita MeSH
- dánio pruhované MeSH
- Daphnia účinky léků MeSH
- ekotoxikologie metody normy MeSH
- elastomery toxicita MeSH
- ethyleny toxicita MeSH
- guma toxicita MeSH
- Heterokontophyta účinky léků MeSH
- odchylka pozorovatele MeSH
- reprodukovatelnost výsledků MeSH
- testy toxicity metody normy MeSH
- vejce MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- chemické látky znečišťující vodu MeSH
- elastomery MeSH
- ethylene-propylene-diene-monomer MeSH Prohlížeč
- ethyleny MeSH
- guma MeSH
