[Identification of Clinical Isolates of the Bacillus cereus Group and Their Characterization by Mass Spectrometry and Electron Microscopy].

Bacillus cereus is a spore-forming bacterium found in the environment mainly in soil. Bacillus spores are known to be extremely resistant not only to environmental factors, but also to various sanitation regimes. This leads to spore contamination of toxin-producing strains in hospital and food equipment and, therefore, poses a great threat to human health. Two clinical isolates identified as B. cereus and B. cytotoxicus were used in the present work. It was shown that their calcium ion content was significantly lower than that of the reference strains. According to electron microscopy, one of the SRCC 19/16 isolates has an enlarged exosporium, and the SRCC 1208 isolate has large electron-dense inclusions of an unclear nature during sporulation. We can assume that these contain a biologically active component with a cytotoxic effect and possibly play a role in pathogenesis. Comparative chemical, biochemical, physiological, and ultrastructural analysis of spores of clinical isolates and reference strains of B. cereus was performed. The results we obtained deepen our understanding of the properties of spores that contribute to the increased pathogenicity of B. cereus group species.

A novel antivirulent compound fluorothiazinone inhibits Klebsiella pneumoniae biofilm in vitro and suppresses model pneumonia.

The problematic treatment of infections caused by multiple-resistant Klebsiella, especially in ICU, is the leading cause of prolonged hospitalization and high mortality rates. The use of antibiotics for the prevention of infections is considered unreasonable as it may contribute to the selection of resistant bacteria. In this regard, the development of drugs that will be effective in preventing infection during various invasive procedures is extremely necessary. We have shown that the developed innovative antibacterial compound fluorothiazinone (FT) that suppresses the formation of biofilms is effective in the prevention of a model pneumonia caused by a multi-resistant clinical K. pneumoniae isolate. Prophylactic use followed by treatment with FT in mice with acute pneumonia modulates the local innate immune response without suppressing protective properties in the early stages of infection, while contributing to a decrease in the bacterial load in the organs and preventing lethal pathological changes in the lungs at later stages of K. pneumoniae infection. Further development of such antivirulence drugs and their use will reduce morbidity and mortality in nosocomial infections, as well as reduce the number of antibiotics used.

Optical and Electron Microscopic Study of the Morphology and Ultrastructure of Biofilms Formed by Streptococcus pyogenes.

Our study confirmed the capacity of S. pyogenes strains to form biofilms on abiotic surfaces. Chains of streptococci surrounded by bluish film were seen under a microscope after alcian blue staining of the preparations grown on slides. On ultrathin sections in transmission electron microscope, the extracellular matrix (indicator of biofilm maturity) became visible after staining with alcian blue. Microscopy of the sections shows structures characteristic of a biofilm in spaces between the cells. Scanning electron microscopy also demonstrates the presence of a biomembrane. Importantly that type 1M strain forming in fact no membranes when cultured on plastic plates (Costar) formed biofilms on the glass. It seems that the conditions for the biofilm formation on the plastic and on the glass differ, due to which the exopolymeric matrices formed on different surfaces vary by biochemical composition.

Ultrastructural organization of Helicobacter pylori under natural conditions and during ex vivo culturing.

Ultrastructure of Helicobacter pylori were studied by transmission electron microscopy in freshly isolated and ex vivo cultured biopsy specimens of gastric antral mucosa from patients with duodenal ulcer.

Ultrastructure of circulating immune complexes isolated from the blood plasma of patients suffering from infectious diseases.

An original procedure which permits to isolate circulating immune complexes (CIC) from the blood plasma in a form of a dense pellet was developed. This procedure was applied for the ultrastructural analysis of CIC isolated from blood of healthy blood donors and patients suffering from Yersinia enterocolitica (YE) and Yersinia pseudotuberculosis (YP) infections. The here reported method of CIC isolation from blood plasma permitted to visualize CIC electronmicroscopically as amorphous masses of low, middle, and high electron density with inclusions of cell debris. In contrast to CIC of healthy blood donors, CIC of infected patients contained various bacteria and fungiformic structures. For the first time, this method made possible an ultrastructural demonstration of bacterial destruction outside of phagocytes in vivo. This method also permits to visualize and identify bacteria in cases of lingering forms of infection when hemoculture tests fail. Therefore, electronmicroscopic examination of CIC preparations from the blood plasma might be a very informative indicator of bacteriemia in the course of an infection process and serve as an indicator of therapeutic effects. In lingering forms of an infection process, ultrastructural analysis of CIC preparations can be of prognostic value and serve as an indicator of therapeutic effects. This method might be also advantageous as an additional test for the exposure of latent bacterial persistence in diagnostically complicated cases.