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.

• Keywords
• Biotechnology, Cyanobacteria, Flow cytometry, Fluorescent viability probes, Stress resistance, Viability assessment,
• MeSH
• Fluorescent Dyes * MeSH
• Stress, Physiological * MeSH
• Microbial Viability * MeSH
• Polyhydroxyalkanoates metabolism   MeSH
• Flow Cytometry * methods   MeSH
• Cyanobacteria * physiology   MeSH
• Synechocystis * physiology   MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Fluorescent Dyes * MeSH
• Polyhydroxyalkanoates MeSH

Phase contrast imaging is well-suited for studying weakly scattering samples. Its strength lies in its ability to measure how the phase of the electron beam is affected by the sample, even when other imaging techniques yield low contrast. In this study, we explore via simulations two phase contrast techniques: integrated center of mass (iCOM) and ptychography, specifically using the extended ptychographical iterative engine (ePIE). We simulate the four-dimensional scanning transmission electron microscopy (4D-STEM) datasets for specific parameters corresponding to a scanning electron microscope (SEM) with an immersive objective and a given pixelated detector. The performance of these phase contrast techniques is analyzed using a contrast transfer function. Simulated datasets from a sample consisting of graphene sheets and carbon nanotubes are used for iCOM and ePIE reconstructions for two aperture sizes and two electron doses. We highlight the influence of aperture size, showing that for a smaller aperture, the radiation dose is spent mostly on larger sample features, which may aid in imaging sensitive samples while minimizing radiation damage.

• Keywords
• 4D-STEM, SEM, STEM, carbon nanotubes, ePIE, electron microscopy, iCOM, low dose, phase contrast, ptychography,
• Publication type
• Journal Article MeSH

Purple photosynthetic bacteria (PPB) are versatile microorganisms capable of producing various value-added chemicals, e.g., biopolymers and biofuels. They employ diverse metabolic pathways, allowing them to adapt to various growth conditions and even extreme environments. Thus, they are ideal organisms for the Next Generation Industrial Biotechnology concept of reducing the risk of contamination by using naturally robust extremophiles. Unfortunately, the potential of PPB for use in biotechnology is hampered by missing knowledge on regulations of their metabolism. Although Rhodospirillum rubrum represents a model purple bacterium studied for polyhydroxyalkanoate and hydrogen production, light/chemical energy conversion, and nitrogen fixation, little is known regarding the regulation of its metabolism at the transcriptomic level. Using RNA sequencing, we compared gene expression during the cultivation utilizing fructose and acetate as substrates in case of the wild-type strain R. rubrum DSM 467T and its knock-out mutant strain that is missing two polyhydroxyalkanoate synthases PhaC1 and PhaC2. During this first genome-wide expression study of R. rubrum, we were able to characterize cultivation-driven transcriptomic changes and to annotate non-coding elements as small RNAs.

• Keywords
• Acetate, Depolymerase knock-out, Fructose, Gene ontology, Genome, Metabolism, Polyhydroxyalkanoates, RNA-Seq, Rhodospirillum rubrum, Transcriptome,
• Publication type
• Journal Article MeSH