Olga Tarasenko

Decontamination of biological warfare agents by a microwave plasma torch

A portable arc-seeded microwave plasma torch running stably with airflow is described and applied for the decontamination of biological warfare agents. Emission spectroscopy of the plasma torch indicated that this torch produced an abundance of reactive atomic oxygen that could effectively oxidize biological agents. Bacillus cereus was chosen as a simulant of Bacillus anthracis spores for biological agent in the decontamination experiments. Decontamination was performed with the airflow rate of 0.393l∕s, corresponding to a maximum concentration of atomic oxygen produced by the torch. The experimental results showed that all spores were killed in less than 8 s at 3 cm distance, 12 s at 4 cm distance, and 16 s at 5 cm distance away from the nozzle of the torch.

Contribution of a portable air plasma torch to rapid blood coagulation as a method of preventing bleeding

The effectiveness and mechanism of a low temperature air plasma torch in clotting blood are explored. Both blood droplets and smeared blood samples were used in the tests. The treated droplet samples reveal how blood clotting depends on the distance at which the torch operated, and for how long the droplets have been exposed to the torch. Microscopy and cell count of smeared blood samples shed light on dependencies of erythrocyte and platelet counts on torch distance and exposure time. With an increase of torch distance, the platelet count of treated blood samples increases but is less than that of the control. The flux of reactive atomic oxygen (RAO) and the degree of blood clotting decreased. With an increase of exposure time, platelet count of treated samples decreased, while the degree of clot increased. The correlation among these dependencies and published data support a blood clotting mechanism that RAO as well as other likely reactive oxygen species generated by the plasma torch activate erythrocyte-platelets interactions and induces blood coagulation.

Plasma effects on bacterial spores in a wet environment

An arc-seed microwave plasma torch, which can run stably at low airflow rate (e.g., 0.393 l s−1) and produces an abundance of reactive atomic oxygen in its plasma effluent, is applied for studying the effects of atomic oxygen on bacterial spores in solution. Bacillus cereus was chosen as the biological agent. The experimental results show that the plasma effluent can penetrate into water to kill B. cereus spores. The kill time (i.e., 10-fold reduction time) is about 10 s at an exposure distance of 3 cm, 24 s at 4 cm, and 31 s at 5 cm. Morphological studies are performed via scanning electron and atomic force microscopes, which take two- and three-dimensional images of spores to record the changes in their morphological structures and shapes caused by the plasma effluent. The loss of appendages and exosporium in the structure as well as flattened cell shapes are observed.

Killing of Bacterial Spores Contained in a Paper Envelope by a Microwave Plasma Torch

The air plasma effluent of an arc-seed microwave torch is used to kill bacterial spores contained inside an envelope. The torch is operated at a 60-Hz periodic mode (with about 40% duty cycle) and runs stably at a low airflow rate (e.g., 0.393 l/s). The images of plasma torch plumes show that the arc loop of the discharge prolongs from the electrodes by nearly 3 cm and microwave energizes the charge particles along the arc loop considerably. The emission spectroscopy of the torch indicates that the plasma effluent contains an abundance of reactive atomic oxygen. Bacillus cereus is chosen for the biological agent in the decontamination experiment. The experiment and the decontamination efficacy of this torch are presented. The averaged temperature inside the envelope is measured to be less than 40degC, thus ruling out the thermal decontamination mechanism

Scanning Electron and Atomic Force Microscopy to Study Plasma Torch Effects on B. cereus Spores

The occurrence of scanning electron microscopy (SEM) and atomic force microscopy (AFM) side-by-side is becoming increasingly common in analytical research. This article shows microscopy techniques to image Bacillus spores, to measure spore dimensions, and to demonstrate how these methods provide supplementary information to study plasma torch effects. This paper demonstrates that observed morphologies of spores before and after exposure to a plasma torch are remarkably different. The use of SEM and AFM as a tool complex enables examination of spore morphology and dimensions as well as their alterations during decontamination using plasma torch

Killing of Bacillus spores is mediated by nitric oxide and nitric oxide synthase during glycoconjugate-enhanced phagocytosis.

Nitric oxide (NO) is a signaling and defense molecule of major importance. NO endows macrophages with bactericidal, cytostatic as well as cytotoxic activity against various pathogens. Bacillus spores can produce serious diseases, which might be attenuated if macrophages were able to kill the spores on contact. Present research was carried out to study whether glycoconjugates stimulated NO and nitric oxide synthase (NOS2) production during phagocytosis killing of Bacillus spores. Murine macrophages exposed to glycoconjugate-treated spores induced NOS2 and NO production that was correlated with high viability of macrophages and killing rate of bacterial spores. Increased levels of inducible NOS2 and NO production by macrophages in presence of glycoconjugates suggested that the latter provide an activation signal directed to macrophages. Glycoconjugates were shown to exert a protective influence, sparing macrophages from spore-induced cell death. In presence of glycoconjugates, macrophages efficiently kill the organisms. Without glycoconjugate activation, murine macrophages were ineffective at killing Bacillus spores. These results suggest that glycoconjugates promote killing of Bacillus spores by blocking spore-induced macrophage cell death, while increasing their activation level and NO and NOS2 production. Glycoconjugates suggest novel antimicrobial approaches to prevention and treatment of infection caused by bacterial spores.

Glycoconjugates enhanced the intracellular killing of Bacillus spores, increasing macrophage viability and activation

Infections caused by Bacillus spores can be attenuated if the intracellular killing of the organism by macrophages can be enhanced. Glycoconjugate-bearing polymers, which selectively bind to Bacillus spores, were tested for modulation of intracellular killing when added prior to, during, and following macrophage exposure to B. cereus spores. In the absence of glycoconjugates, murine macrophages were ineffective at killing Bacillus spores. In presence of glycoconjugates, however, macrophages efficiently killed spores, whether the glycoconjugates were added to the cells prior to, during, and following spore addition. Glycoconjugates were shown to exert a protective influence on macrophages and increase their activation, as evidenced by viability and lactate dehydrogenase release assays. Increased levels of nitric oxide production by macrophages pretreated with glycoconjugates suggest that, under these conditions, glycoconjugates provide an activation signal to macrophages. These results indicate that glycoconjugates promote killing of Bacillus spores, while increasing macrophage activation level and viability. The selection of glycoconjugate ligands bearing immunomodulating properties could be exploited for vaccine and/or immunomodulator development and/or for the improvement of existing vaccines against B. cereus and B. anthracis.

Fan-shaped microwave plasma for mail decontamination

A microwave torch is designed to produce fan-shaped plasma, which extends about 140 mm laterally. This torch produces an abundance of reactive atomic oxygen in the plasma effluent as evidenced by its emission spectroscopy. The results of the spectral intensity measurements show that the produced atomic oxygen outside the microwave cavity distributes quite uniformly over a width of about 80 mm and reaches out more than 10 mm. An experiment applying this plasma to kill Bacillus cereus contained in an envelope has been performed. The kill rate is presented.

Polymeric glycoconjugates protect and activate macrophages to promote killing of Bacillus cereus spores during phagocytosis

Diseases caused by Bacillus spores might be attenuated if macrophages were able to kill the spores on exposure. Glycoconjugate-bearing polymers, which have been shown to bind to Bacillus spores, were tested for modulation of phagocytosis of B. cereus spores. Without glycoconjugate activation, murine macrophages were ineffective at killing Bacillus spores during phagocytosis. In the presence of glycoconjugates, however, the macrophages efficiently killed the organisms. The glycoconjugates were shown to have a protective influence, sparing macrophages from spore-induced cell death. Very low concentrations of the glycoconjugates prevented macrophage cell death, as shown by lactate dehydrogenase (LDH) release and trypan blue assays. Increased levels of inducible nitric oxide (NO) production by the macrophages in the presence of glycoconjugates suggested that the glycoconjugates provide an activation signal to the macrophages. These results suggest that glycoconjugates promote the killing of Bacillus spores by blocking spore-induced macrophage cell death, while increasing their activation level. Polymeric glycoconjugates may suggest novel approaches to improve existing vaccines as well as prevent and treat infections incurred through either B. cereus or B. anthracis spores.

Instrumental improvements and sample preparations that enable reproducible, reliable acquisition of mass spectra from whole bacterial cells

Rationale Rapid sub-species characterization of pathogens is required for timely responses in outbreak situations. Pyrolysis mass spectrometry (PyMS) has the potential to be used for this purpose. Methods However, in order to make PyMS practical for traceback applications, certain improvements related to spectrum reproducibility and data acquisition speed were required. The main objectives of this study were to facilitate fast detection (<30 min to analyze 6 samples, including preparation) and sub-species-level bacterial characterization based on pattern recognition of mass spectral fingerprints acquired from whole cells volatilized and ionized at atmospheric pressure. An AccuTOF DART mass spectrometer was re-engineered to permit ionization of low-volatility bacteria by means of Plasma Jet Ionization (PJI), in which an electric discharge, and, by extension, a plasma beam, impinges on sample cells. Results Instrumental improvements and spectral acquisition methodology are described. Performance of the re-engineered system was assessed using a small challenge set comprised of assorted bacterial isolates differing in identity by varying amounts. In general, the spectral patterns obtained allowed differentiation of all samples tested, including those of the same genus and species but different serotypes. Conclusions Fluctuations of ±15% in bacterial cell concentrations did not substantially compromise replicate spectra reproducibility.