A collection of 120 strains isolated from stool specimens collected from humans suffering from gastroenteritis and from environmental samples were analyzed by random amplified polymorphic DNA PCR (RAPD), repetitive extragenic palindromic PCR (REP-PCR), and enterobacterial repetitive intergenic consensus sequence PCR (ERIC-PCR). Species of Aeromonoas hydrophila, A. bestiarum, A. salmonicida, A. caviae, A. media, and A. veronii revealed clonal structure. There was no dominant clone causing gastroenteritis in humans. Moreover, there was no genetic similarity between clinical and environmental strains of Aeromonas sp. isolated from different geographical areas as well as from the same geographical area. Some clones colonized specific ecosystems, e.g., drinking water distribution systems. RAPD and ERIC-PCR methods had the same discriminatory power and proved to be useful for epidemiological investigation and population genetic analysis of Aeromonas spp., whereas REP-PCR was less effective for differentiating the isolates of Aeromonas spp.Bacteria of Aeromonas sp. are gram-negative, straight cells (rod-shaped to coccoid) with rounded ends. They are oxidase and catalase positive, reduce nitrate to nitrite, and ferment D-glucose. These bacteria are widely spread in the environment, especially in surface water and sewage; they also occur in untreated and treated drinking water (1, 2, 4, 18). In humans, Aeromonas spp. are responsible for gastroenteritis, chronic diarrhea, wound infections, respiratory tract infections, peritonitis, urinary tract infections, and septicemia (2, 17). Among Aeromonas-associated infections of humans A. hydrophila, A. caviae, and A. veronii are the predominating species, whereas A. eucrenophila, A. popoffii, (2), and A. culicicola (28) have never been found in clinical samples. Some Aeromonas species are associated with a wide variety of diseases in cold-and warm-blooded animals, including fish, frogs, water buffaloes, reptiles, birds, and cattle (16, 28). A. veronii, A. hydrophila, and A. salmonicida are capable of causing septicemia in freshwater and marine fish (3, 9). Clinical and environmental Aeromonas sp. isolates secrete many extracellular products, such as hemolysins, enterotoxins, and proteases. Studies conducted by Kühn et al. (22) showed that some isolates of a given species produce virulence factors more frequently than others. These findings indicate that the virulence within the genus Aeromonas might be a clonal property and only some clones may be responsible for progressive disease. However, there have been no studies that would have determined clonal structure within Aeromonas spp. and the spread of specific clones in human population and in the environment. Our study was undertaken to recognize the clonal relatedness of strains derived from diarrheal stool specimens collected from humans living in different geographical areas. We also compared genetic similarities of clones recovered from stool and the environment. In addition, we attempted to determine the genetic relati...

Coagulase-negative staphylococci (CoNS) are opportunistic pathogens that particularly cause infections in patients with implanted medical devices. The present research was performed to study the virulence potential of 53 clinical isolates of Staphylococcus capitis, Staphylococcus auricularis, Staphylococcus lugdunensis, Staphylococcus simulans, Staphylococcus cohnii and Staphylococcus caprae. All clinical strains were clonally unrelated. Isolates carried genes encoding resistance to β-lactam (mecA) (15 %), aminoglycoside [aac(6')/aph(2″)(11 %), aph (3')-IIIa (15 %), ant(4')-Ia (19 %)] and macrolide, lincosamide and streptogramin B (MLSB) [erm(A) (4 %), erm(B) (13 %), erm(C) (41 %), msr(A) (11 %)] antibiotics. CoNS isolates (64 %) were able to form biofilms. Confocal laser scanning microscopy revealed that these biofilms formed a three-dimensional structure composed mainly of living cells. All biofilm-positive strains carried the ica operon. In vitro studies demonstrated that a combination treatment with tigecycline and rifampicin was more effective against biofilms than one with ciprofloxacin and rifampicin. The minimum biofilm eradication concentration values were 0.062-0.5 µg ml-1 for tigecycline/rifampicin and 0.250-2 µg ml-1 for ciprofloxacin/rifampicin. All CoNS strains adhered to the human epithelial cell line HeLa, and more than half of the isolates were able to invade the HeLa cells, although most invaded relatively poorly. The virulence of CoNS is also attributed to their cytotoxic effects on HeLa cells. Incubation of HeLa cells with culture supernatant of the CoNS isolates resulted in cell death. The results indicate that the pathogenicity of S. capitis, S. auricularis, S. lugdunensis, S. cohnii and S. caprae is multi-factorial, involving the ability of these bacteria to adhere to human epithelial cells, form biofilms and invade and destroy human cells.

Staphylococcus hominis is the third species of coagulase-negative staphylococci (CoNS) most frequently isolated from specimens of patients with hospital-acquired infections. Many infections caused by CoNS appeared to be associated with biofilms. Nevertheless, the knowledge of the ability of S. hominis to form a biofilm is limited. The aim of this study was to analyze the formation of the biofilm by 56 S. hominis strains isolated from clinical cases. The biofilm three-dimensional structure was reconstructed by confocal laser scanning microscopy. We found that most of S. hominis strains carried icaADBC genes encoding polysaccharide intercellular adhesin (PIA), which plays a crucial role in the formation of biofilms in staphylococci strains. However, only a half of the ica-positive strains had an ability to form a biofilm in vitro. In this study, we also accessed the sensitivity of biofilms of S. hominis strains to sodium metaperiodate, proteinase K and DNase. We found that polysaccharides and proteins are the major components of the extracellular matrix of the biofilm formed by S. hominis. DNase did not have a significant effect on biofilms, which suggested that nucleic acid plays a minor role in the mature biofilm.

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The aim of this study was to investigate the interaction of Staphylococcus haemolyticus strains with a macrophage cell line. Infection with the strains resulted in macrophage injury. All strains exhibited cytotoxic effects towards J774 cells. Moreover, the bacteria triggered apoptosis of the cells. The lowest apoptotic index did not exceed 21 %, whereas the highest reached 70 % at 24 h and 85 % at 48 h after infection. Incubation with the bacteria caused loss of mitochondrial membrane potential (ΔΨm) in macrophages. The pro-apoptotic activity of the strains was blocked by a pan-caspase inhibitor z-VAD-fmk, indicating the involvement of caspases in the bacteria-mediated cell death. We observed that the induction of macrophage apoptosis could constitute an important mechanism of pathogenesis by which S. haemolyticus strains evade host immune defences and cause disease.