Genomic diversity of two lineages of exfoliative toxin A-converting phages predominating in Staphylococcus aureus strains in the Czech Republic.

We have isolated and characterized two distinct types of exfoliative toxin A (ETA)-converting bacteriophages originating from Staphylococcus aureus strains responsible for massive outbreaks of pemphigus neonatorum in the Czech Republic. Three induced phages designated as ph iB531, phi B557 and phi B122 were found to be capable of transferring the eta gene into the prophageless non-toxigenic S. aureus strain and converting it into an ETA producer. Comparisons of the phage sequences derived from 12 selected genes and 2 genomic segments (polymorphic P2 and conserved C4) revealed that phi B531 and phi B557 were identical each other, but phi B122 differed from them in 5 gene sequences, the xis gene content and the virion protein profile. Thus, phi B122 represents a new type of still undescribed ETA-converting phage. This study highlights not only the conclusive genomic diversity of eta gene-positive phages, but also their virulence implications in impetigo S. aureus strains.

Photochemical properties of a potential photosensitiser indocyanine green in vitro.

The present study investigates the photochemical properties of a potential photosensitiser, indocyanine green (ICG), in an in vitro HeLa cell system. Cell proliferation was studied after a combined effect of ICG, at a concentration range of 24-94 microM, and therapeutic laser irradiation at several different energy densities. In addition, ICG cytotoxicity was evaluated in HeLa cells and V79 Chinese hamster by the MTT assay. Phototoxicity was evaluated at 1, 24, and 48 h after irradiation. No phototoxic effect was detected 1h after irradiation. The maximum phototoxic effect of ICG on HeLa cells was detected for an ICG concentration of 94 microM, a laser output of 360 mW, and an energy density of 99 J/cm(2) at 24h after irradiation. Potentiation of the ICG phototoxic effect was achieved by adding 20 microM H(2)O(2), which was a non-toxic concentration for HeLa cells in this experimental design. At 48 h after laser irradiation a statistically significant difference was found between the toxicity of ICG plus peroxide, as compared to ICG alone. The addition of H(2)O(2) at a concentration of 20 microM caused a significant increase in phototoxicity of ICG for HeLa cells. Our results confirm that ICG could be a perspective agent for use in photodynamic therapy and that its phototoxic effect can be potentiated by addition of an oxidative agent.

Discrete Fourier transform-based analysis of HeLa cell microtubules after ultrasonic exposure.

Cytoskeletal structures can be affected by external factors including ultrasound. Our task was to develop a structure analysis method to evaluate these changes quantitatively. We exposed HeLa cells to continuous ultrasound (1 MHz, 1 and 2 W/cm2, 10 min at 37 degrees C). The microtubules were detected by the monoclonal antibody TU-01/SwAM/FITC, observed in a fluorescence microscope and photographed digitally. The images were processed by "FFT magic" software. The structure analysis is based on frequency domain filtering using discrete Fourier transform. The basic idea is to design filters to extract information describing best the structural changes. The properties of the filter can be enhanced by direction filtering, i.e., extraction of a symmetric angular segment in the frequency domain centered on a zero frequency. The final image is a normalized sum of inverse FFT's of such segmented spectra. We needed a method yielding a single number assigned to the structure, e.g., the ratio of the area of microtubules to the total cell area. Assuming that the image background intensity is constant, we can use thresholding to detect areas occupied by the cells. The information about the area of the microtubules is contained in a wide range of higher intensities. Therefore, we use a gamma correction. The area occupied by microtubules is then considered an area with intensities above the selected threshold. There were tested three different filters to extract information about microtubules. The mathematical method chosen seems sensitive enough for quantitative assessment of changes of the microtubular network.

Effects of low-power laser irradiation on cell locomotion in protozoa.

Low-power lasers are commonly used in human medicine for treatment of various pathological conditions, but mechanisms of their healing effects are still poorly understood. The results of this study provide information related to these effects at the cellular level. Two different protozoan species, Euglena gracilis and Tetrahymena thermophila, were used to study changes in locomotion behavior in response to low-power lasers. The cells were irradiated at 830 and 650 nm generated by a semiconductor laser (99 J/cm2, 360 mW) and a laser pointer (0.75 J/cm2, 5 mW), respectively, and their locomotion was recorded by a TV camera and analyzed using computer software. Exposure to laser light, regardless of the wavelength, resulted in increased cell velocity in both species (P <0.001). Exposure to 650 nm produced an equal increase in median cell velocity in both E. gracilis (19.0%) and T. thermophila (18.2%), and some increase persisted in the postirradiation 30 s period. Irradiation by the 830 nm laser resulted in a markedly higher response in Tetrahymena (29.4%) than in Euglena (15.2%), and the two median values remained increased after irradiation was discontinued. Different reactions found in the species studied and some mechanisms underlying the response of cells to radiation are discussed.