This study introduces an evaluation methodology tailored for bioreactors, with the aim of assessing the stress experienced by algae due to harmful contaminants released from antifouling (AF) paints. We present an online monitoring system equipped with an ultra-sensitive sensor that conducts non-invasive measurements of algal culture's optical density and physiological stage through chlorophyll fluorescence signals. By coupling the ultra-sensitive sensor with flash-induced chlorophyll fluorescence, we examined the dynamic fluorescence changes in the green microalga Chlamydomonas reinhardtii when exposed to biocides. Over a 24-h observation period, increasing concentrations of biocides led to a decrease in photosynthetic activity. Notably, a substantial reduction in the maximum quantum yield of primary photochemistry (FV/FM) was observed within the first hour of exposure. Subsequently, we detected a partial recovery in FV/FM; however, this recovery remained 50% lower than that of the controls. Integrating the advanced submersible sensor with fluorescence decay kinetics offered a comprehensive perspective on the dynamic alterations in algal cells under the exposure to biocides released from antifouling coatings. The analysis of fluorescence relaxation kinetics revealed a significant shortening of the fast and middle phases, along with an increase in the duration of the slow phase, for the coating with the highest levels of biocides. Combining automated culturing and measuring methods, this approach has demonstrated its effectiveness as an ultrasensitive and non-invasive tool for monitoring the physiology of photosynthetic cultures. This is particularly valuable in the context of studying microalgae and their early responses to various environmental conditions, as well as the potential to develop an AF system with minimal harm to the environment.

• Klíčová slova
• Antifouling coatings, Bioreactor, Chlorophyll fluorescence spectroscopy, Microalgae, Toxicity, Ultrasensitive sensor,
• MeSH
• bioreaktory * MeSH
• chemické látky znečišťující vodu analýza   MeSH
• Chlamydomonas reinhardtii * účinky léků   metabolismus   MeSH
• chlorofyl metabolismus   MeSH
• dezinficiencia farmakologie   MeSH
• fluorescence MeSH
• fotosyntéza účinky léků   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• chemické látky znečišťující vodu MeSH
• chlorofyl MeSH
• dezinficiencia MeSH

Although microalgae have only recently been recognized as part of the plant and soil microbiome, their application as biofertilizers has a tradition in sustainable crop production. Under consideration of their ability to produce the plant growth-stimulating hormone cytokinin (CK), known to also induce pathogen resistance, we have assessed the biocontrol ability of CK-producing microalgae. All pro- and eukaryotic CK-producing microalgae tested were able to enhance the tolerance of tobacco against Pseudomonas syringae pv. tabaci (PsT) infection. Since Chlamydomonas reinhardtii (Cre) proved to be the most efficient, we functionally characterized its biocontrol ability. We employed the CRISPR-Cas9 system to generate the first knockouts of CK biosynthetic genes in microalgae. Specifically, we targeted Cre Lonely Guy (LOG) and isopentenyltransferase (IPT) genes, the key genes of CK biosynthesis. While Cre wild-type exhibits a strong protection, the CK-deficient mutants have a reduced ability to induce plant defence. The degree of protection correlates with the CK levels, with the IPT mutants showing less protection than the LOG mutants. Gene expression analyses showed that Cre strongly stimulates tobacco resistance through defence gene priming. This study functionally verifies that Cre primes defence responses with CK, which contributes to the robustness of the effect. This work contributes to elucidate microalgae-mediated plant defence priming and identifies the role of CKs. In addition, these results underscore the potential of CK-producing microalgae as biologicals in agriculture by combining biofertilizer and biocontrol ability for sustainable and environment-friendly crop management.

• MeSH
• Chlamydomonas reinhardtii * genetika   metabolismus   MeSH
• CRISPR-Cas systémy * MeSH
• cytokininy * metabolismus   MeSH
• mutace MeSH
• nemoci rostlin * mikrobiologie   imunologie   genetika   MeSH
• odolnost vůči nemocem * genetika   MeSH
• Pseudomonas syringae patogenita   fyziologie   MeSH
• tabák * genetika   mikrobiologie   imunologie   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• cytokininy * MeSH

β-Lactamases (EC 3.5.2.6) confer resistance against β-lactam group-containing antibiotics in bacteria and higher eukaryotes, including humans. Pathogenic bacterial resistance against β-lactam antibiotics is a primary concern for potential therapeutic developments and drug targets. Here, we report putative β-lactamase activity, sulbactam binding (a β-lactam analogue) in the low μM affinity range, and site-specific interaction studies of a 14 kDa UV- and dark-inducible protein (abbreviated as UVI31+, a BolA homologue) from Chlamydomonas reinhartii. Intriguingly, the solution NMR structure of UVI31 + bears no resemblance to other known β-lactamases; however, the sulbactam binding is found at two sites rich in positively charged residues, mainly at the L2 loop regions and the N-terminus. Using NMR spectroscopy, ITC and MD simulations, we map the ligand binding sites in UVI31 + providing atomic-level insights into its β-lactamase activity. Current study is the first report on β-lactamase activity of UVI31+, a BolA analogue, from C. reinhartii. Furthermore, our mutation studies reveal that the active site serine-55 is crucial for β-lactamase activity.

• Klíčová slova
• ITC, MD, NMR, S55A, Sulbactam, UVI31+, β-lactam antibiotics, β-lactamases,
• MeSH
• beta-laktamasy * chemie   metabolismus   MeSH
• Chlamydomonas reinhardtii * enzymologie   MeSH
• magnetická rezonanční spektroskopie metody   MeSH
• nukleární magnetická rezonance biomolekulární metody   MeSH
• rostlinné proteiny chemie   metabolismus   MeSH
• sekvence aminokyselin MeSH
• simulace molekulární dynamiky MeSH
• sulbaktam chemie   farmakologie   MeSH
• vazba proteinů MeSH
• vazebná místa MeSH
• Publikační typ
• časopisecké články MeSH

Demonstrating outdoor cultivation of engineered microalgae at considerable scales is essential for their prospective large-scale deployment. Hence, this study focuses on the outdoor cultivation of an engineered Chlamydomonas reinhardtii strain, 3XAgBs-SQs, for bisabolene production under natural dynamic conditions of light and temperature. Our preliminary outdoor experiments showed improved growth, but frequent culture collapses in conventional Tris-acetate-phosphate medium. In contrast, modified high-salt medium (HSM) supported prolonged cell survival, outdoor. However, their subsequent outdoor scale-up from 250 mL to 5 L in HSM was effective with 10 g/L bicarbonate supplementation. Pulse amplitude modulation fluorometry and metabolomic analysis further validated their improved photosynthesis and uncompromised metabolic fluxes towards the biomass and the products (natural carotenoids and engineered bisabolene). These strains could produce 906 mg/L bisabolene and 54 mg/L carotenoids, demonstrating the first successful outdoor photoautotrophic cultivation of engineeredC. reinhardtii,establishing it as a one-cell two-wells biorefinery.

• Klíčová slova
• Algal bio-refinery, Bicarbonate supplementation, Carotenoids, Growth engineering, Natural dynamic conditions,
• MeSH
• Chlamydomonas reinhardtii * metabolismus   MeSH
• Chlamydomonas * metabolismus   MeSH
• fotosyntéza MeSH
• karotenoidy metabolismus   MeSH
• prospektivní studie MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• karotenoidy MeSH

Two-dimensional materials have recently gained significant awareness. A representative of such materials, black phosphorous (BP), earned attention based on its comprehensive application potential. The presented study focuses on the mode of cellular response underlying the BP interaction with Chlamydomonas reinhardtii as an algal model organism. We observed noticeable ROS formation and changes in outer cellular topology after 72 h of incubation at 5 mg/L BP. Transcriptome profiling was employed to examine C. reinhardtii response after exposure to 25 mg/L BP for a deeper understanding of the associated processes. The RNA sequencing has revealed a comprehensive response with abundant transcript downregulation. The mode of action was attributed to cell wall disruption, ROS elevation, and chloroplast disturbance. Besides many other dysregulated genes, the cell response involved the downregulation of GH9 and gametolysin within a cell wall, pointing to a shift to discrete manipulation with resources. The response also included altered expression of the PRDA1 gene associated with redox governance in chloroplasts implying ROS disharmony. Altered expression of the Cre-miR906-3p, Cre-miR910, and Cre-miR914 pointed to those as potential markers in stress response studies.

• Klíčová slova
• 2D material, BP, Cre-miR910, Cre-miR914, MiRNAs Cre-miR906–3p, RNAseq, Toxicity,
• MeSH
• Chlamydomonas reinhardtii * metabolismus   MeSH
• chloroplasty genetika   metabolismus   MeSH
• fosfor metabolismus   MeSH
• pochopení MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• transkriptom MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fosfor MeSH
• reaktivní formy kyslíku MeSH

The use of unicellular algae to remove xenobiotics (including drugs) from wastewaters is one of the rapidly developing areas of environmental protection. Numerous data indicate that for efficient phycoremediation three processes are important, i.e. biosorption, bioaccumulation, and biotransformation. Although biosorption and bioaccumulation do not raise any serious doubts, biotransformation is more problematic since its products can be potentially more toxic than the parent compounds posing a threat to organisms living in a given environment, including organisms that made this transformation. Thus, two questions need to be answered before the proper algae strain is chosen for phycoremediation, namely what metabolites are produced during biotransformation, and how resistant is the analyzed strain to a mixture of parent compound and metabolites that appear over the course of culture? In this work, we evaluated the remediation potential of the model green alga Chlamydomonas reinhardtii in relation to non-steroidal anti-inflammatory drugs (NSAIDs), as exemplified by diclofenac. To achieve this, we analysed the susceptibility of C. reinhardtii to diclofenac as well as its capability to biosorption, bioaccumulation, and biotransformation of the drug. We have found that even at a relatively high concentration of diclofenac the algae maintained their vitality and were able to remove (37.7%) DCF from the environment. A wide range of phase I and II metabolites of diclofenac (38 transformation products) was discovered, with many of them characteristic rather for animal and bacterial biochemical pathways than for plant metabolism. Due to such a large number of detected products, 18 of which were not previously reported, the proposed scheme of diclofenac transformation by C. reinhardtii not only significantly contributes to broadening the knowledge in this field, but also allows to suggest possible pathways of degradation of xenobiotics with a similar structure. It is worth pointing out that a decrease in the level of diclofenac in the media observed in this study cannot be fully explained by biotransformation (8.4%). The mass balance analysis indicates that other processes (total 22%), such as biosorption, a non-extractable residue formation, or complete decomposition in metabolic cycles can be involved in the diclofenac disappearance, and those findings open the prospects of further research.

• Klíčová slova
• Biotransformation, Chlamydomonas reinhardtii, Diclofenac, Non-steroidal anti-inflammatory drug,
• MeSH
• antiflogistika nesteroidní analýza   MeSH
• biotransformace MeSH
• chemické látky znečišťující vodu * analýza   MeSH
• Chlamydomonas reinhardtii * metabolismus   MeSH
• diklofenak toxicita   metabolismus   MeSH
• voda MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antiflogistika nesteroidní MeSH
• chemické látky znečišťující vodu * MeSH
• diklofenak MeSH
• voda MeSH

Graphene oxides (GOs) and their reduced forms are often discussed both positively and negatively due to the lack of information about their chemistry and structure. This study utilized GOs with two sheet sizes that were further reduced by two reducing agents (sodium borohydride and hydrazine) to obtain two different degrees of reduction. The synthesized nanomaterials were characterized using scanning electron microscopy (SEM), atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), elemental analysis (EA), Fourier transform infrared (FTIR) spectroscopy, and Raman spectroscopy (RA) to understand their chemistry and structure. The second focus of our investigation included in vitro testing of the biocompatibility/toxicity of these materials on a model organism, the freshwater microalga Chlamydomonas reinhardtii. The effects were studied on the basis of biological endpoints complemented by biomass investigation (FTIR spectroscopy, EA, and atomic absorption spectrometry (AAS)). The results showed that the biocompatibility/toxicity of GOs is dependent on their chemistry and structure and that it is impossible to generalize the toxicity of graphene-based nanomaterials.

• Klíčová slova
• Biocompatibility, Chlamydomonas, Graphene oxide, Microalgae, Reduction, Toxicity,
• MeSH
• Chlamydomonas reinhardtii * MeSH
• grafit * toxicita   MeSH
• nanostruktury * MeSH
• oxidy toxicita   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• grafit * MeSH
• graphene oxide MeSH Prohlížeč
• oxidy MeSH

Recently, more accessible transcriptomic approaches have provided a new and deeper understanding of environmental toxicity. The present study focuses on the transcriptomic profiles of green microalgae Chlamydomonas reinhardtii exposed to new industrially promising material, TiO2 nanotubes (NTs), as an example of a widely used one-dimensional nanomaterial. The first algal in vitro assay included 2.5 and 7.5 mg/L TiO2 NTs, resulting in a dose-dependent negative effect on biological endpoints. At a working concentration of 7.5 mg/L, RNA-sequencing showed a mainly negative effect on the cells. In summary, the results indicated metabolic disruption, such as ATP loss, damage to mitochondria and chloroplasts, loss of solutes due to permeated membranes, and cell wall damage. Moreover, apoptosis-induced transcripts were detected. Interestingly, reactivation of transposons was observed. In signalling and transcription pathways, including chromatin remodelling and locking, the annotated genes were downregulated.

• Klíčová slova
• Chlamydomonas, Nanotubes, RNA-seq, TiO(2), Toxicity,
• MeSH
• chemické látky znečišťující vodu * toxicita   MeSH
• Chlamydomonas reinhardtii * MeSH
• nanotrubičky * MeSH
• transkriptom MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• chemické látky znečišťující vodu * MeSH
• titanium dioxide MeSH Prohlížeč

Light is the essential energy source for autotrophically growing organisms, including microalgae. Both light intensity and light quality affect cell growth and biomass composition. Here we used three green algae-Chlamydomonas reinhardtii, Desmodesmus quadricauda, and Parachlorella kessleri-to study the effects of different light intensities and light spectra on their growth. Cultures were grown at three different light intensities (100, 250, and 500 µmol m-2 s-1) and three different light sources: fluorescent lamps, RGB LEDs, and white LEDs. Cultures of Desmodesmus quadricauda and Parachlorella kessleri were saturated at 250 µmol m-2 s-1, and further increasing the light intensity did not improve their growth. Chlamydomonas reinhardtii cultures did not reach saturation under the conditions used. All species usually divide into more than two daughter cells by a mechanism called multiple fission. Increasing light intensity resulted in an increase in maximum cell size and division into more daughter cells. In Parachlorella kessleri cells, the concentration of photosynthetic pigments decreased with light intensity. Different light sources had no effect on algal growth or photosynthetic pigments. The results show a species-specific response of algae to light intensity and support the use of any white light source for their cultivation without negative effects on growth.

• Klíčová slova
• Chlamydomonas reinhardtii, Desmodesmus quadricauda, LED, Parachlorella kessleri, cell growth, fluorescent tube, light intensity,
• MeSH
• biomasa MeSH
• Chlamydomonas reinhardtii * MeSH
• Chlorophyta * MeSH
• fotosyntéza MeSH
• mikrořasy * MeSH
• světlo MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The intensive use of insecticides in global agricultural production has attracted much attention due to its many adverse effects on human health and the environment. In recent years, the utilization of nanotechnology has emerged as a tool to overcome these adverse effects. The aim of this work was to test different microparticles (zinc oxide (ZnO MPs) and silicon dioxide microparticles (SiO2 MPs)), and silver nanoparticles (Ag NPs) and to study their toxicity on a model organism, Tenebrio molitor. A comprehensive comparative study, which included more than a thousand mealworms divided into nine separate groups, was conducted. In addition to pure nano/microparticle solutions, the effect of particles mixed with the microalgae extract Chlamydomonas reinhardtii was also observed. Pure Ag NPs and SiO2 MPs resulted in larval mortality of more than 70% compared to that of pure ZnO MPs, in which the mortality rate was approximately 33%. A mixture of the algal extract with zinc oxide microparticles resulted in mortality that was double compared to that observed with pure ZnO MPs. In parallel, atomic absorption spectrometry (AAS) was used to determine the difference in the concentration of trace elements in the bodies of dead and live larvae.

• MeSH
• Chlamydomonas reinhardtii chemie   MeSH
• insekticidy chemie   toxicita   MeSH
• kovové nanočástice chemie   MeSH
• rostlinné extrakty chemie   MeSH
• stříbro chemie   MeSH
• Tenebrio účinky léků   MeSH
• zinek chemie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• insekticidy MeSH
• rostlinné extrakty MeSH
• stříbro MeSH
• zinek MeSH