L-Forms of Pseudomonas aeruginosa

L-forms of a strain of Pseudomonas aeruginosa were produced by serial subculture of the bacterial form on agar medium containing sucrose as an osmotic stabilizer and carbenicillin. L-forms eventually became stable, i.e., would not revert in the absence of antibiotic, and were adapted to grow well in broth with the osmotic stabilizer. Gross morphology and light microscopic colony morphology were typical of an L-form. L-form cells were approximately spherical and bounded in part by a plasma membrane; they lacked the triple-layer cell wall structure and coarse, electron-dense nucleoidal granules of the parent bacterial form. The L-form, but not the bacterial form, contained cores, organelles previously reported only in group D streptococci. Antibiotic disc-sensitivity studies showed the stable L-form to be as sensitive as, or more sensitive than, the bacterial form to most antibiotics. Exceptions were polymyxin B, colimycin sulfate, and gentamicin, which were more active against the bacterial form. The remainder of the aminoglycosides and cell wall-active antibiotics showed no inhibition of either form. The L-form was more susceptible to cidal activity of normal human serum than the parent form. The L-form exhibited fewer biochemical activities than the parent bacteria or bacterial forms derived by reversion at a time when the L-form was still unstable. L-form colonies appeared colorless, and chemical analysis demonstrated that, if the L-form produces pigment at all, which was not demonstrated, it could not have been more than 3.6% of that produced by the bacterial form.     

Aeruginocine typing of Pseudomonas aeruginosa

Aeruginocine typing using eight indicator strains of Wahba on Bacto tryptone-glucose yeast extract Agar (Difco) with an incubation temperature of 32°C was found to be a reproducible and easy method which could be readily adopted by a diagnostic clinical laboratory. The results of aeruginocine typing of 1500 strains of Ps. aeruginosa suggest that hospitals differ in the range and patterns of aeruginocine types of strains isolated in them. In addition to representatives of nine of the 10 pyocine types of Wahba, a large number of strains isolated in India were found to fall into other groups not described in Wahba's system. Hence, 20 new inhibition patterns (aeruginocine types) are proposed.     

Pyocine typing of Pseudomonas aeruginosa

The growth-inhibitory effect of a small group of different pyocines has been used to differentiate Ps. aeruginosa isolates. A method is described which can be carried out with ordinary media and is easy for routine application. One hundred and one isolates of Ps. aeruginosa were differentiated by four pyocines into 10 pyocine patterns.     

Silver against Pseudomonas aeruginosa biofilms

Silver has been recognized for its antimicrobial properties for centuries. Most studies on the antibacterial efficacy of silver, with particular emphasis on wound healing, have been performed on planktonic bacteria. Our recent studies, however, strongly suggest that colonization of wounds involves bacteria in both the planktonic and biofilm modes of growth. The action of silver on mature in vitro biofilms of Pseudomonas aeruginosa, a primary pathogen of chronic infected wounds, was investigated. The results show that silver is very effective against mature biofilms of P. aeruginosa, but that the silver concentration is important. A concentration of 5-10 mug/mL silver sulfadiazine eradicated the biofilm whereas a lower concentration (1 mug/mL) had no effect. The bactericidal concentration of silver required to eradicate the bacterial biofilm was 10-100 times higher than that used to eradicate planktonic bacteria. These observations strongly indicate that the concentration of silver in currently available wound dressings is much too low for treatment of chronic biofilm wounds. It is suggested that clinicians and manufacturers of the said wound dressings consider whether they are treating wounds primarily colonized either by biofilm-forming or planktonic bacteria.     

Immunological nonidentity of Pseudomonas paucimobilis with Pseudomonas aeruginosa and Pseudomonas cepacia.

This investigation determined the serum agglutination activity and serum bactericidal response after rabbit immunization with Pseudomonas paucimobilis. Agglutination activity of antisera showed a twofold increase in titer from before immunization to 4 weeks post-immunization and peaked at 8 weeks post-immunization with a titer of 1:512. 2-Mercaptoethanol reduction of immunoglobulin M decreased agglutination titers. No major antigens were found to be common from crude antigen preparations of P. paucimobilis, Pseudomonas aeruginosa and Pseudomonas cepacia when tested with antisera to P. paucimobilis. Serum bactericidal activity was found in post-immunization antisera at 8 and 12 weeks against P. paucimobilis, with no activity present before immunization or at 4 weeks post-immunization. Antisera against P. paucimobilis showed no bactericidal activity against P. aeruginosa or P. cepacia.     

Cross-protection by Pseudomonas aeruginosa polysaccharides.

High-molecular-weight polysaccharide from Pseudomonas aeruginosa immunotypes 1 and 2 gave cross-protection in outbred CD-1 mice challenged with the heterologous immunotype organism. Both active immunization with 50 micrograms of polysaccharide, as well as passive transfer of immune serum were effective. The basis for this cross-protection is the ability of high doses of polysaccharide to induce antibody formation to both homologous and heterologous immunotype determinants.     

Azithromycin retards Pseudomonas aeruginosa biofilm formation

Using a flow cell biofilm model, we showed that a sub-MIC of azithromycin (AZM) can delay but not inhibit Pseudomonas aeruginosa biofilm formation and results in the development of a stable AZM resistance phenotype. Furthermore, mature biofilms were not affected by AZM.     

Physiologie der Nitratreduktion bei Pseudomonas aeruginosa

By both Pseudomonas strains the N₂ and N₂O production depended on the “physiological condition” of the bacteria. The N₂O formation by strain C 31 in mineral media depended also on the precultivation medium. The bacteria produced N₂O besides N₂, if precultivated in Nutrient Broth + KNO₃. Precultivated in a mineral medium, however, they produced only N₂. The anaerobic gas production of Pseudomonas aeruginosa strain C 31 and strain N.C.I.B 8704 was determined under hydrogen atmosphere. The results are as follows: In Nutrient Broth + 0.35% KNO₃ nitrate was respirated quantitatively to N₂ and N₂O by the two strains. On the contrary, in Nutrient Broth + 0.5% KNO₃ nitrate was not completely consumed by strain C 31. In nitrate-ammonium-medium we succeeded to recover quantitatively the given nitrogen compounds (nitrate, nitrite, ammonium, organic N), partially in the denitrification products, after the end of the experiment, but not in the experiment with nitrate-medium. In nitrate-medium strain C 31 produced a much higher amount of nitrite (up to 73 mg N/l) as in nitrate-ammonium-medium (up to 213 μg N/l). The influence of the ammonium ions is discussed.     

Polysaccharide antigens of Pseudomonas aeruginosa.

The major polysaccharide antigens of P. aeruginosa are the cell-wall lipopolysaccharides many of which have an acidic polysaccharide chain (O-antigen) rich in unusual amino sugars. The D-rhamnose-rich polysaccharide antigen common to many serologically distinct strains is also associated with the lipopolysaccharide. The high-molecular-weight polysaccharides with O-specificity are present in extracellular slime produced by strains isolated from the environmental and from the immunocompromised hosts. The extracellular antigenic polysaccharide of another type (bacterial alginate) is expressed by mucoid strains isolated from patients with cystic fibrosis. Serotype-specific immune responses after infection are directed at the lipopolysaccharides and these heat-stable antigens serve as the basis for differentiation of P. aeruginosa strains. Both the cell-wall antigens including conjugates of the O-polysaccharides with different proteins and the extracellular antigens have been used to prepare specific antibodies tested for protection against infections due to P. aeruginosa.     

Complement activation by Pseudomonas aeruginosa biofilms

In chronic infections, such as the bronchopulmonary Pseudomonas aeruginosa infection in cystic fibrosis (CF) patients, bacteria persist despite an intact host immune defense and frequent antibiotic treatment. An important reason for the persistence of the bacteria is their capacity for the biofilm mode of growth. In this study we investigated the role of biofilms in activation of complement, a major contributor to the inflammatory process. Complement activation by P. aeruginosa was examined in a complement consumption assay, production of C3 and factor B conversion products assessed by crossed immuno-electrophoresis, C5a generation tested by a PMN chemotactic assay, and terminal complement complex formation measured by ELISA. Two of the four assays showed that P. aeruginosa grown in biofilm activated complement less than planktonic bacteria, and all assays showed that activation by intact biofilms was submaximal. Factor B conversion was of low magnitude indicating the importance of the classical pathway. Complement activation by P. aeruginosa was inhibited by polymyxin B indicating that lipopolysaccharide (LPS) was the main mediator of complement activation. Immune complexes and massive influx of neutrophils are known to cause inflammatory changes in the lungs. P. aeruginosa persisting in biofilms may contribute to the constant inflammation taking place in the lungs of CF patients.     

Further isolations of non-flagellate Pseudomonas aeruginosa

Three strains of non-flagellate Ps. aeruginosa isolated from pathological specimens have already been described. The purpose of this paper is to report further isolates. All strains have been shown to be different types isolated from a variety of infections.