Effect of toluene on Pseudomonas stutzeri ST-9 morphology — Plasmolysis, cell size, and formation of outer membrane vesicles

Isolated toluene-degrading Pseudomonas stutzeri ST-9 bacteria were grown in a minimal medium containing toluene (100 mg·L⁻¹) (MMT) or glucose (MMG) as the sole carbon source, with specific growth rates of 0.019 h⁻¹ and 0.042 h⁻¹, respectively. Scanning (SEM) as well as transmission (TEM) electron microscope analyses showed that the bacterial cells grown to mid-log phase in the presence of toluene possess a plasmolysis space.TEManalysis revealed that bacterial cells that were grown in MMT were surrounded by an additional “material” with small vesicles in between. Membrane integrity was analyzed by leakage of 260 nm absorbing material and demonstrated only 7% and 8% leakage from cultures grown in MMT compared with MMG. X-ray microanalysis showed a 4.3-fold increase in Mg and a 3-fold increase in P in cells grown in MMT compared with cells grown in MMG. Fluorescence-activated cell sorting (FACS) analysis indicated that the permeability of the membrane to propidium iodide was 12.6% and 19.6% when the cultures were grown inMMGand MMT, respectively. The bacterial cell length increased by 8.5% ± 0.1% and 17% ± 2%, as measured using SEM images and FACS analysis, respectively. The results obtained in this research show that the presence of toluene led to morphology changes, such as plasmolysis, cell size, and formation of outer membrane vesicles. However, it does not cause significant damage to membrane integrity.