Search results for: Marketa Vaculovicova

Authors: Simona Dostalova, Dita Munzova, Ana Maria Jimenez Jimenez, Marketa Vaculovicova, Vojtech Adam, Rene Kizek

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The boom of nanomedicine in recent years has led to the development of numerous new nanomaterials that can be used as nanocarriers in the drug delivery. These nanocarriers can either be synthetic or natural-based. The disadvantage of many synthetic nanocarriers is their toxicity in patient’s body. Protein cages that can naturally be found in human body do not exhibit such disadvantage. However, the release of cargo from some protein cages in target cells can be problematic. As a special type of protein cages can serve the capsid of many viruses, including phage. Phages infect bacterial cells; therefore they are not harmful to human cells. The targeting of phage particles to cancer cells can be solved by producing of empty phage capsids during which the targeting moieties (e.g. peptides) can be cloned into genes of phage capsid to decorate its surface. Moreover, the produced capsids do not contain viral nucleic acid and are therefore not infectious to beneficial bacteria in the patient’s body. The protein cage composed of viral capsid is larger than other frequently used apoferritin cage but its size is still small enough to benefit from passive targeting by Enhanced Permeability and Retention effect. In this work, bacteriophage λ was used, both whole and its empty capsid for delivery of different cytotoxic drugs (cisplatin, carboplatin, oxaliplatin, etoposide and doxorubicin). Large quantities of phage λ were obtained from phage λ-producing strain of E. coli cultivated in medium with 0.2 % maltose. After killing of E. coli with chloroform and its removal by centrifugation, the phage was concentrated by ultracentrifugation at 130 000 g and 4 °C for 3 h. The encapsulation of the drugs was performed by infusion method and four different concentrations of the drugs were encapsulated (200; 100; 50; 25 µg/ml). Free molecules of drugs were removed by dialysis. The encapsulation was verified using spectrophotometric and electrochemical methods. The amount of encapsulated drug linearly increased with the amount of applied drug (determination coefficient R2=0.8013). 76% of applied drug was encapsulated in phage λ particles (concentration of 10 µg/ml), even with the highest applied concentration of drugs, 200 µg/ml. Only 1% of encapsulated drug was detected in phage DNA. Similar results were obtained with encapsulation in phage empty capsid. Therefore, it can be concluded that the encapsulation of drugs into phage particles is efficient and mostly occurs by interaction of drugs with protein capsid.

Keywords: cytostatics, drug delivery, nanocarriers, phage capsid

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Authors: Iva Blazkova, Amitava Moulick, Vedran Milosavljevic, Pavel Kopel, Marketa Vaculovicova, Vojtech Adam, Rene Kizek

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Doxorubicin is one of the most commonly used and the most effective chemotherapeutic drugs. This antracycline drug isolated from the bacteria Streptomyces peuceticus var. caesius is sold under the trade name Adriamycin (hydroxydaunomycin, hydroxydaunorubicin). Doxorubicin is used in single therapy to treat hematological malignancies (blood cancers, leukaemia, lymphoma), many types of carcinoma (solid tumors) and soft tissue sarcomas. It has many serious side effects like nausea and vomiting, hair lost, myelosupression, oral mucositis, skin reactions and redness, but the most serious one is the cardiotoxicity. Because of the risk of heart attack and congestive heart failure, the total dose administered to patients has to be accurately monitored. With the aim to lower the side effects and to targeted delivery of doxorubicin into the tumor tissue, the different nanoparticles are studied. The drug can be bound on a surface of nanoparticle, encapsulated in the inner cavity, or incorporated into the structure of nanoparticle. Among others, carbon nanoparticles (graphene, carbon nanotubes, fullerenes) are highly studied. Besides the number of inorganic nanoparticles, a great potential exhibit also organic ones mainly lipid-based and polymeric nanoparticle. The aim of this work was to perform a toxicity study of free doxorubicin compared to doxorubicin conjugated with various nanotransporters. The effect of liposomes, fullerenes, graphene, and carbon nanotubes on the toxicity was analyzed. As a first step, the binding efficacy of between doxorubicin and the nanotransporter was determined. The highest efficacy was detected in case of liposomes (85% of applied drug was encapsulated) followed by graphene, carbon nanotubes and fullerenes. For the toxicological studies, the chicken embryos incubated under controlled conditions (37.5 °C, 45% rH, rotation every 2 hours) were used. In 7th developmental day of chicken embryos doxorubicin or doxorubicin-nanotransporter complex was applied on the chorioallantoic membrane of the eggs and the viability was analyzed every day till the 17th developmental day. Then the embryos were extracted from the shell and the distribution of doxorubicin in the body was analyzed by measurement of organs extracts using laser induce fluorescence detection. The chicken embryo mortality caused by free doxorubicin (30%) was significantly lowered by using the conjugation with nanomaterials. The highest accumulation of doxorubicin and doxorubicin nanotransporter complexes was observed in the liver tissue

Keywords: doxorubicin, chicken embryos, nanotransporters, toxicity

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Authors: Simona Dostalova, Pavel Kopel, Marketa Vaculovicova, Vojtech Adam, Rene Kizek

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Protein apoferritin seems to be a very promising structure for use as a nanocarrier. It is prepared from intracellular ferritin protein naturally found in most organisms. The role of ferritin proteins is to store and transport ferrous ions. Apoferritin is a hollow protein cage without ferrous ions that can be prepared from ferritin by reduction with thioglycolic acid or dithionite. The structure of apoferritin is composed of 24 protein subunits, creating a sphere with 12 nm in diameter. The inner cavity has a diameter of 8 nm. The drug encapsulation process is based on the response of apoferritin structure to the pH changes of surrounding solution. In low pH, apoferritin is disassembled into individual subunits and its structure is “opened”. It can then be mixed with any desired cytotoxic drug and after adjustment of pH back to neutral the subunits are reconnected again and the drug is encapsulated within the apoferritin particles. Excess drug molecules can be removed by dialysis. The receptors for apoferritin, SCARA5 and TfR1 can be found in the membrane of both healthy and cancer cells. To enhance the specific targeting of apoferritin nanocarrier, it is possible to modify its surface with targeting moieties, such as antibodies. To ensure sterically correct complex, we used a a peptide linker based on a protein G with N-terminus affinity towards Fc region of antibodies. To connect the peptide to the surface of apoferritin, the C-terminus of peptide was made of cysteine with affinity to gold. The surface of apoferritin with encapsulated doxorubicin (ApoDox) was coated either with gold nanoparticles (ApoDox-Nano) or gold (III) chloride hydrate reduced with sodium borohydride (ApoDox-HAu). The applied amount of gold in form of gold (III) chloride hydrate was 10 times higher than in the case of gold nanoparticles. However, after removal of the excess unbound ions by electrophoretic separation, the concentration of gold on the surface of apoferritin was only 6 times higher for ApoDox-HAu in comparison with ApoDox-Nano. Moreover, the reduction with sodium borohydride caused a loss of doxorubicin fluorescent properties (excitation maximum at 480 nm with emission maximum at 600 nm) and thus its biological activity. Fluorescent properties of ApoDox-Nano were similar to the unmodified ApoDox, therefore it was more suited for the intended use. To evaluate the specificity of apoferritin modified with antibodies, we used ELISA-like method with the surface of microtitration plate wells coated by the antigen (goat anti-human IgG antibodies). To these wells, we applied ApoDox without targeting antibodies and ApoDox-Nano modified with targeting antibodies (human IgG antibodies). The amount of unmodified ApoDox on antigen after incubation and subsequent rinsing with water was 5 times lower than in the case of ApoDox-Nano modified with targeting antibodies. The modification of non-gold ApoDox with antibodies caused no change in its targeting properties. It can therefore be concluded that the demonstrated procedure allows us to create nanocarrier with enhanced targeting properties, suitable for nanomedicine.

Keywords: apoferritin, doxorubicin, nanocarrier, targeting antibodies

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Authors: Markéta Gregušová, Vladimíra Schindlerová, Ivana Šajdlerová, Petr Mohyla, Jan Kedroň

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Company managers are always looking for more and more opportunities to succeed in today's fiercely competitive market. To maintain your place among the successful companies on the market today or to come up with a revolutionary business idea is much more difficult than before. Each new or improved method, tool, or approach that can improve the functioning of business processes or even of the entire system is worth checking and verification. The use of simulation in the design of manufacturing systems and their management in practice is one of the ways without increased risk, which makes it possible to find the optimal parameters of manufacturing processes and systems. The paper presents an example of use of simulation for solution of the bottleneck problem in the concrete company.

Keywords: practical applications, business processes, systems, simulation

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Authors: Marketa Kotova, Jana Kolarova, Ludek Zalud, Petr Dobsak

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A game for training of breath (TRABR) for continuous monitoring of pulmonary ventilation during the patients’ therapy focuses especially on monitoring of their ventilation processes. It is necessary to detect, monitor and differentiate abdominal and thoracic breathing during the therapy. It is a fun form of rehabilitation where the patient plays and also practicing isolated breathing. Finally the game to practice breath was designed to evaluate whether the patient uses two types of breathing or not.

Keywords: pulmonary ventilation, thoracic breathing, abdominal breathing, breath monitoring using pressure sensors, game TRABR TRAining of BReath)

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Authors: Martina Perinkova, Lenka Kolarcikova, Marketa Twrda

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In Ostrava there are a lot of former post-industrial areas and areas that have gradually through conversions and their subsequent reuse. One of the largest is the national cultural monument Lower Vítkovice area where there is a large complex transformation of the former iron production. Industrial heritage today visited by tourists for entertainment, culture, history, sports and other activities. This is a unique example of reuse of technical monuments and introduction of new life into the historic area. The main task of not only find the right function and use, in terms of re integration into city life and finding a balance between history and current lifestyle, looking at the history of the area and its technical condition before reconstruction. It is not only very expensive but also time consuming. Transformations industrial monument is the result of a dialogue architect, the idea of the investor and expert opinion heritage institute.

Keywords: post-industrial area, cultural monument, conversions

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Authors: Marketa Slazanska

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Cicero is generally perceived as one of the most gifted authors of all the antiquity, which has brought him a great popularity. Even the orator himself was eager to be seen as a man of fine taste for culture and especially literature, which was in his days represented by the interest in everything of a Greek origin. It might, therefore, lead to an assumption that literature was a key interest in his life, too. However, a confirmation of this assumption cannot be found in his letters, which are believed to offer Cicero's most personal statements. Reading his letters thoroughly and comparing them to the respective historical background, it may be found out that the author has mentioned literature mainly in a specific context of his political career losing importance. To analyze the letters have been used in the original Latin version as well as several translations and a large historical and literary background. The object of this paper is to specify, whether Cicero regarded himself more as an author and orator, or as a politician, and how did his correspondents regard him. The respective findings could be useful for better understanding of author's life and work and they may help his readers to see his books from a different, and perhaps more accurate, point of view.

Keywords: antiquity, politician, literature, letters, Cicero, oratory

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Authors: Markéta Slováková

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The largest share of policy and money within the European Union goes to agriculture. The Union’s Common Agricultural Policy has undergone several transformations in the last five decades, with the main change taking place in the 1990's. This change influenced agriculture in the Czech Republic, inasmuch as the fledgling republic was preparing to join the European Union and adopt its policies. In the 1990s, Czech agriculture passed from a centrally planned economy to a market economy and subsequently adopted the terms of the Common Agricultural Policy. The Czech Republic is also characterized by a significant landscape sphere diversification. Agricultural entrepreneurs in the Czech Republic are still not used the possibility of grants from the European Union. They focus rather on national or regional subsidy titles. Only half of all agricultural entrepreneurs in the Czech Republic use European subsidies. This article focuses on the introduction of the Common Agricultural Policy to the Czech Republic and its subsequent influence on Czech agriculture. It is demonstrated on the implementation rate of the CAP in the EU Member States and the closer focus is on the Czech integration.

Keywords: common agricultural policy, agriculture, European Union, transformation

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Authors: Martina Perinkova, Lenka Kolarcikova, Marketa Twrda

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Brownfields are one of the most important problems that must be solved by today's cities. The topic of this article is description of developing a comprehensive transformation of post-industrial area of the former iron factory national cultural heritage Lower Vítkovice. City of Ostrava used to be industrial superpower of the Czechoslovak Republic, especially in the area of coal mining and iron production, after declining industrial production and mining in the 80s left many unused areas of former factories generally brownfields and backfields. Since the late 90s we are observing how the city officials or private entities seeking to remedy this situation. Regeneration of brownfields is a very expensive and long-term process. The area is now rebuilt for tourists and residents of the city in the entertainment, cultural, and social center. It was necessary do the reconstruction of the industrial monuments. Equally important was the construction of new buildings, which helped reusing of the entire complex. This is a unique example of transformation of technical monuments and completion of necessary new objects, so that the area could start working again and reintegrate back into the urban system.

Keywords: brown fields, conversion, historical and industrial buildings, reconstruction

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Authors: Olena Mayboroda, Angel Gonzalez Benito, Jonathan Sabate Del Rio, Marketa Svobodova, Sandra Julich, Herbert Tomaso, Ciara K. O'Sullivan, Ioanis Katakis

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DNA amplification is required for most molecular diagnostic applications but conventional PCR has disadvantages for field testing. Isothermal amplification techniques are being developed to respond to this problem. One of them is the Recombinase Polymerase Amplification (RPA) that operates at isothermal conditions without sacrificing specificity and sensitivity in easy-to-use formats. In this work RPA was used for the optical detection of solid-phase amplification of the potential biowarfare agent Yersinia pestis. Thiolated forward primers were immobilized on the surface of maleimide-activated microtitre plates for the quantitative detection of synthetic and genomic DNA, with elongation occurring only in the presence of the specific template DNA and solution phase reverse primers. Quantitative detection was achieved via the use of biotinylated reverse primers and post-amplification addition of streptavidin-HRP conjugate. The overall time of amplification and detection was less than 1 hour at a constant temperature of 37oC. Single-stranded and double-stranded DNA sequences were detected achieving detection limits of 4.04*10-13 M and 3.14*10-16 M, respectively. The system demonstrated high specificity with negligible responses to non-specific targets.

Keywords: recombinase polymerase amplification, Yersinia pestis, solid-phase detection, ELONA

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Authors: Maria Stepankova, Lucie Vistejnova, Pavel Klein, Tereza Blassova, Marketa Slajerova, Radek Sedlacek, Martin Bartos, Jaroslav Chlupac

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Background: Clinical outcome benefits of the segment rib fracture surgical therapy are well known and follow from better stabilization of the chest wall. Despite this, some authors still incline to conservative therapy and point out to possible rib fracture healing failure in connection with the bone vascular supply disturbance caused by metal plate implantation. This suggestion met neither experimental nor clinical verification and remains the object of discussion. In our pilot study we investigated the titanium plate fixation effect on the rib fracture healing in porcine model and its histological, biomechanical and radiological aspects. Materials and Method: Two porcine models (experimental group) underwent the operative chest wall stabilization with a titanium plate implantation after osteotomy. Two other porcine models (control group) were treated conservatively after osteotomy. Three weeks after surgery, all animals were sacrificed, treated ribs were explanted and the histological analysis, µCT imaging and biomechanical testing of the calluses tissue were performed. Results: In µCT imaging, experimental group showed a higher cortical bone volume compared to the control group. Histological analysis using the non-decalcified bone tissue blocks demonstrated more maturated callus with higher newly-formed osseous tissue ratio in experimental group in comparison to controls. In contrast, no significant differences in bone blood vessels supply in both groups were observed. This finding suggests that the bone blood supply in experimental group was not impaired. Biomechanical analysis using 3-point bending test demonstrated significantly higher bending stiffness and the maximum force in experimental group. Conclusion: Based on our observation, it could be concluded, that the titanium plate fixation of the rib fractures leads to faster bone callus maturation whereas does not cause the vascular supply impairment after 3 weeks and thus has a beneficial effect on the rib fracture healing.

Keywords: bone vascular supply, chest wall stabilization, fracture healing, histological analysis, titanium plate implantation

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Authors: Miroslav Mrlik, Marketa Ilcikova, Martin Cvek, Josef Osicka, Michal Sedlacik, Vladimir Pavlinek, Jaroslav Mosnacek

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Magnetorheological elastomers (MREs) are a unique type of materials consisting of two components, magnetic filler, and elastomeric matrix. Their properties can be tailored upon application of an external magnetic field strength. In this case, the change of the viscoelastic properties (viscoelastic moduli, complex viscosity) are influenced by two crucial factors. The first one is magnetic performance of the particles and the second one is off-state stiffness of the elastomeric matrix. The former factor strongly depends on the intended applications; however general rule is that higher magnetic performance of the particles provides higher MR performance of the MRE. Since magnetic particles possess low stability properties against temperature and acidic environment, several methods how to improve these drawbacks have been developed. In the most cases, the preparation of the core-shell structures was employed as a suitable method for preservation of the magnetic particles against thermal and chemical oxidations. However, if the shell material is not single-layer substance, but polymer material, the magnetic performance is significantly suppressed, due to the in situ polymerization technique, when it is very difficult to control the polymerization rate and the polymer shell is too thick. The second factor is the off-state stiffness of the elastomeric matrix. Since the MR effectivity is calculated as the relative value of the elastic modulus upon magnetic field application divided by elastic modulus in the absence of the external field, also the tuneability of the cross-linking reaction is highly desired. Therefore, this study is focused on the controllable modification of magnetic particles using a novel monomeric system based on 2-(1H-pyrrol-1-yl)ethyl methacrylate. In this case, the short polymer chains of different chain lengths and low polydispersity index will be prepared, and thus tailorable stability properties can be achieved. Since the relatively thin polymer chains will be grafted on the surface of magnetic particles, their magnetic performance will be affected only slightly. Furthermore, also the cross-linking density will be affected, due to the presence of the short polymer chains. From the application point of view, such MREs can be utilized for, magneto-resistors, piezoresistors or pressure sensors especially, when the conducting shell on the magnetic particles will be created. Therefore, the selection of the pyrrole-based monomer is very crucial and controllably thin layer of conducting polymer can be prepared. Finally, such composite particle consisting of magnetic core and conducting shell dispersed in elastomeric matrix can find also the utilization in shielding application of electromagnetic waves.

Keywords: atom transfer radical polymerization, core-shell, particle modification, electromagnetic waves shielding

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Authors: Markéta Chmelíková, David Ullrich, Iva Burešová

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The paper maps the possibilities and limits of diagnosing selected personality and performance characteristics of military leadership and psychology students in the context of coping with challenging situations. Individuals vary greatly inter-individually in their ability to effectively manage extreme situations, yet existing diagnostic tools are often criticized mainly for their low predictive power. Nowadays, every modern army focuses primarily on the systematic minimization of potential risks, including the prediction of desirable forms of behavior and the performance of military commanders. The context of military leadership is well known for its life-threatening nature. Therefore, it is crucial to research stress load in the specific context of military leadership for the purpose of possible anticipation of human failure in managing extreme situations of military leadership. The aim of the submitted pilot study, using an experiment of 24 hours duration, is to verify the possibilities of a specific combination of psychodiagnostic to predict people who possess suitable equipment for coping with increased stress load. In our pilot study, we conducted an experiment of 24 hours duration with an experimental group (N=13) in the bomb shelter and a control group (N=11) in a classroom. Both groups were represented by military leadership students (N=11) and psychology students (N=13). Both groups were equalized in terms of study type and gender. Participants were administered the following test battery of personality characteristics: Big Five Inventory 2 (BFI-2), Short Dark Triad (SD-3), Emotion Regulation Questionnaire (ERQ), Fatigue Severity Scale (FSS), and Impulsive Behavior Scale (UPPS-P). This test battery was administered only once at the beginning of the experiment. Along with this, they were administered a test battery consisting of the Test of Attention (d2) and the Bourdon test four times overall with 6 hours ranges. To better simulate an extreme situation – we tried to induce sleep deprivation - participants were required to try not to fall asleep throughout the experiment. Despite the assumption that a stay in an underground bomb shelter will manifest in impaired cognitive performance, this expectation has been significantly confirmed in only one measurement, which can be interpreted as marginal in the context of multiple testing. This finding is a fundamental insight into the issue of stress management in extreme situations, which is crucial for effective military leadership. The results suggest that a 24-hour stay in a shelter, together with sleep deprivation, does not seem to simulate sufficient stress for an individual, which would be reflected in the level of cognitive performance. In the context of these findings, it would be interesting in future to extend the diagnostic battery with physiological indicators of stress, such as: heart rate, stress score, physical stress, mental stress ect.

Keywords: bomb shelter, extreme situation, military leadership, psychodiagnostic

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Authors: Jan Masak, Eva Kvasnickova, Vladimir Scholtz, Olga Matatkova, Marketa Valkova, Alena Cejkova

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Microbial colonization of medical instruments, catheters, implants, etc. is a serious problem in the spread of nosocomial infections. Biofilms exhibit enormous resistance to environment. The resistance of biofilm populations to antibiotic or biocides often increases by two to three orders of magnitude in comparison with suspension populations. Subjects of interests are substances or physical processes that primarily cause the destruction of biofilm, while the released cells can be killed by existing antibiotics. In addition, agents that do not have a strong lethal effect do not cause such a significant selection pressure to further enhance resistance. Non-thermal plasma (NTP) is defined as neutral, ionized gas composed of particles (photons, electrons, positive and negative ions, free radicals and excited or non-excited molecules) which are in permanent interaction. In this work, the effect of NTP generated by the cometary corona with a metallic grid on the formation and stability of biofilm and metabolic activity of cells in biofilm was studied. NTP was applied on biofilm populations of Staphylococcus epidermidis DBM 3179, Pseudomonas aeruginosa DBM 3081, DBM 3777, ATCC 15442 and ATCC 10145, Escherichia coli DBM 3125 and Candida albicans DBM 2164 grown on solid media on Petri dishes and on the titanium alloy (Ti6Al4V) surface used for the production joint replacements. Erythromycin (for S. epidermidis), polymyxin B (for E. coli and P. aeruginosa), amphotericin B (for C. albicans) and ceftazidime (for P. aeruginosa) were used to study the combined effect of NTP and antibiotics. Biofilms were quantified by crystal violet assay. Metabolic activity of the cells in biofilm was measured using MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) colorimetric test based on the reduction of MTT into formazan by the dehydrogenase system of living cells. Fluorescence microscopy was applied to visualize the biofilm on the surface of the titanium alloy; SYTO 13 was used as a fluorescence probe to stain cells in the biofilm. It has been shown that biofilm populations of all studied microorganisms are very sensitive to the type of used NTP. The inhibition zone of biofilm recorded after 60 minutes exposure to NTP exceeded 20 cm², except P. aeruginosa DBM 3777 and ATCC 10145, where it was about 9 cm². Also metabolic activity of cells in biofilm differed for individual microbial strains. High sensitivity to NTP was observed in S. epidermidis, in which the metabolic activity of biofilm decreased after 30 minutes of NTP exposure to 15% and after 60 minutes to 1%. Conversely, the metabolic activity of cells of C. albicans decreased to 53% after 30 minutes of NTP exposure. Nevertheless, this result can be considered very good. Suitable combinations of exposure time of NTP and the concentration of antibiotic achieved in most cases a remarkable synergic effect on the reduction of the metabolic activity of the cells of the biofilm. For example, in the case of P. aeruginosa DBM 3777, a combination of 30 minutes of NTP with 1 mg/l of ceftazidime resulted in a decrease metabolic activity below 4%.

Keywords: anti-biofilm activity, antibiotic, non-thermal plasma, opportunistic pathogens

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