Ultrastructural Aspects of Autolysis of Pseudomonas fluorescens Induced by Osmotic Shock

Autolysis of exponentially growing Pseudomonas fluorescens was induced by a rapid treatment with distilled water before treatment with 0·5 m-sodium acetate, pH 6·5, and incubation for 2 h at 30 °C. Electron microscopy revealed three stages of autolysis. In the first stage most of the cells were plasmolysed. In the second stage (from 5 to 20 min) the number of plasmolysed cells diminished, the profile of the outer membrane changed from asymmetric to symmetric, fractures appeared in the outer membrane and signs of peptidoglycan degradation became apparent. In the third stage (which began after 20 to 60 min) the ultrastructural alterations of the cell wall became more serious, the cytoplasmic membrane exhibited micromorphological lesions and progressive cellular disintegration occurred. The addition of chloramphenicol (100 μg ml−1) or tetracycline (4·5 μg ml−1), to either the culture or the suspensions of autolysing cells, did not significantly reduce the rate of autolysis. The presence of Ca2+ or Mg2+ (10 mM) in the autolytic medium inhibited autolysis and there were no significant ultrastructural alterations. The presence of 2 mm-N -bromosuccinimide (an inhibitor of bacterial autolytic enzymes) in the autolytic medium strongly inhibited autolysis of P. fluorescens;however, the bacteria incubated under these conditions showed significant ultrastructural alterations (extensive splitting of, and occasional fractures in, the outer membrane; cytoplasmic membrane and ribosomes not visible).